CHEMICAL EFFECTS ON WATER-USEEFFICIENCY OF ALFALFA (MEDICAGO SATIVA L.)

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Authors Cole, Darrell Franklin, 1941-

Publisher The University of Arizona.

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COLE, Darrell Franklin, 1941-CHEMICAL EFFECTS ON WATER-USE EFFICIENCY OF ALFALFA (MEDICAGO SATIVA L.).

University of Arizona, Ph.D., 1971 Agronomy

University Microfilms, A XEROX Company, Ann Arbor, Michigan

THIS DISSERTATION HAS BEEN MICROFILMED EXACTLY AS RECEIVED

CHEMICAL EFFECTS' ON WATER-USE EFFICIENCY OF

ALFALFA (MED.ICAGO SATIVA L. )

by

Darrell Franklin Cole

A Dissertation Submitted to the Faculty of the

DEPARTMENT OF AGRONOMY

In Partial Fulfillment of the Requirements For the Degree of

DOCTOR OF PHILOSOPHY

In the Graduate College

THE UNIVERSITY OF ARIZONA

19 7 1

THE UNIVERSITY OF ARIZONA

GRADUATE COLLEGE

I hereby recommend that this dissertation prepared under my

direction by Parrel 1 Franklin Cole

entitled Chemical Effects on Water-use Efficiency of Alfalfa

(Medicaqo sat iva L.)•

be accepted as fulfilling the dissertation requirement of the

degree of Doctor of Philosophy

^ J < * / / £ / 7 # Dissertation Director £T~ Date '

After inspection of the final copy of the dissertation, the

following members of the Final Examination Committee concur in

its approval and recommend its acceptance:*

L. . A*' j'- u. i /•/• *w / / -A/" / ./ / l ,

''' S • <• v /a /n J~> o:

/a/z/r*

(Qikxt £. /0/7/pD 7 /

This approval and acceptance is contingent on the candidate's adequate performance and defense of this dissertation at the final oral examination. The inclusion of this sheet bound into the library copy of the dissertation is evidence of satisfactory performance at the final examination.

STATEMENT BY AUTHOR

This dissertation has been submitted in partial fulfillment of requirements for an advanced degree at The University of Arizona and is deposited in the University Library to be made available to borrowers under rules of the Library.

Brief quotations from this dissertation are allowable without special permission, provided that accurate acknowledgment of source is made. Requests for permission for extended quotation from or reproduction of this manuscript in whole or in part may be granted by the head of the major department or the Dean of the Graduate College when in his judgment the proposed use of the material is in the interests of scholarship. In all other instances, however, permission must be obtained from the author.

SIGNED:

ACKNOWLEDGMENTS

The author is deeply indebted to Dr. Albert K.

Dobrenz for his constructive advice, suggestions, and his

untiring assistance throughout the course of graduate

study.

Acknowledgnient is given to the graduate committee,

Dr. Martin A. Massengale, Dr. L. Nea.1 Wright, Dir. Waltei"

S. Phillips, and Dr. Robert S. Mellor, for their guidance

and assistance in preparation of this dissertation.

The author also wants to thank Dr. L. Neal Wright

and Dr. Gerald 1-1. Luper for providing greenhouse and

growth chamber facilities.

Acknowledgment is given to the Department of

Agronomy for providing monetary funds to assist the author

in his graduate program.

The author expresses his gi-atitude to his wife and

children for their encouragement and patience during the

graduate program.

iii

TADLE OF CONTENTS

Page

LIST OF ILLUSTRATIONS v

LIST OF TABLES viii

ABSTRACT xi

INTRODUCTION 1

REVIEW OF LITERATURE 3

Water Requirement of Alfalfa 3 Effect of Chemicals on Water-Use

Efficiency 4 Transpiration Rates of Alfalfa 5 Effect of Chemicals on Transpiration 6 Photosynthetic Rates of Alfalfa 8 Effect of Chemicals on Photosynthesis

and Respiration ........ 9 Effect of Chemicals on Alfalfa 11

MATERIALS AND METHODS 12

Lath and Greenhouse Experiments 13 Growth Chamber Experiments l6

Transpiration 17 Photosynthesis and Respiration 20

Field Experiment 20

RESULTS AND DISCUSSION . . . 25

Lathhouse and Greenhouse Experiments 25 Growth Chamber Experiments 34

Effect of GA on Transpii-ation 44 Effect of GA on Photosynthesis

and Respiration 49 Field Experiments 62

SUMMARY 83

REFERENCES 86

iv

LIST OF ILLUSTRATIONS

Figure Page

1. Plexiglass chamber used to measure the photosynthetic rates of entire alfalfa plants. Note the air intake near the base and exit near the top. The temperature probe is shown in the middle of the chamber 19

2. Plexiglass chamber used to measure photosynthetic and respiration rates of plants grown in the field 23

3> Effect of GA on length of individual internodes of alfa.l.fa grown in a controlled environment. Internodes arc numbered consecutively from cutoff level (one) to stem tip (ten) 40

4. Effect of GA 011 height of alfalfa stems grown in a controlled environment 4l

5- Effect of GA on the rate of stem elongation in alfalfa grown in a controlled environment 42

6. Transpiration rates for two clones of Mesa-Sirsa alfalfa grown .in a controlled environment 45

7. Effect of GA on the transpiration rates of Mesa-Sirsa alfalfa grown in a contro1Ied environment 47

8. Effect of GA on the transpiration rates o f t wo c. lone s o f M e s a - S i r s a a I f < 1 1. f a grown in a controlled environment 48

9. Effect of GA 011 the percentage moisture and total forage of Mesa-Sirsa alfalfa grown in a cont rolled environment 52

10. Effect of GA 011 the stem-petiole and leaflet, weights of Mesa-Sirsa alfalfa grown in a controlled environment 53

v

vi

LIST OF ILLUSTRATIONS--Continued

Figure Page

11. Effect of GA on transpired water and water requirement of Mesa-Sirsa alfalfa grown in a controlled environment 5 4

12. Effect of GA on leaflet to stem-petiole ratio and specific leaf weight of Mesa-Sirsa alfalfa grown in a controlled environment 55

13• Effect of GA on respiration rates of Mesa-Sirsa alfalfa grown in a controlled environment. Left) ing CO^ dm~2 hr-1. Right) nig ^2 ®

l4. Effect of GA on pliotosyntlietic rates of Mesa-Sirsa alfalfa grown in a controlled environment. Left) mg CO2 dm~2 hr-1. Right) mg COg g--*- hr-1 57

15* Effect of GA on the yield of Mesa-Sirsa alfalfa grown under field conditions. Data taken from an area of .19 rn^ 63

l6. Effect of GA on stem length and secondary branching. l) Control. 2) Treated .... 66

17* Effect of GA on alfalfa leaf anatomy. Note the increased width find the arrangement of palisade and spongy mesophyll cells (1250 X). Upper) Treated. Lower) Control- 69

18. Effect of GA on raceme and sepal elonga­tion. l) Control. 2) Treated 71

19. Effect of GA on photosynthesis (mg CO2 dm~2 hr-1) of alfalfa grown under field conditions 72

20. Effect of GA on respiration (mg CO2 dm_2 hr-1) of alfalfa grown under field conditions 73

21. Effect of GA on specific leaf weight (SLW) of alfalfa grown under field conditions 7'j

vii

LIST OF ILLUSTRATIONS--Continued

Figure Page

22. Chlorosis and increased height of Mesa-Sirsa alfalfa following GA application under field conditions 76

23. Effect of GA on per cent trarismittance of a chlorophyll extract of Mesa-Sirsa alfalfa 79

2'i. Photograph showing the separation of free sugars by thin layer chroiiuitography. Standards were prepared at a concentra­tion of 2 nig/ml . Maltose was spotted with 10 and 25 |i.l , Mj_ and M2, respec­tively. Sucrose (S) and fructose (F) were spotted with 25 jJ,l and the unknown was spotted with 5, 10, 25, and 50 jxl , la, lb, 1c , and Id, respectively 8l

LIST OF TABLES

Table Page

1. Chemicals, rates, and methods of application used on seeded plants of Mesa-Sirs a alfalfa in a controlled greenhouse environment l'l

2. Chemicals and rates used on seeded plants of Mesa-Sirsa alfalfa in a lathhouse environment 15

3- Effect of various chemicals on water-use efficiency of Mesa-Sirsa alfalfa grown in a greenhouse environment 26

k. Effect of various chemicals on stem-petiole and leaflet weight of Mesa-Sirsa alfalfa grown in a greenhouse environment 27

5. Effect of various chemicals on total forage per plant and leaflet to steni-petiole ratio of Mesa-Sirsa alfalfa grown in a greenhouse environment 29

6. Means of water-use efficiency, total forage, and stem-petiole tissue as affected by various chemicals applied to Mesa-Sirsa alfalfa grown in a lathhouse environment at the Tucson Plant Materials Center 31

7. Means of leaflet to stem-petiole ratios, height, and transpired wat er as affected by various chemicals applied to Mesa-Sir Set a I l'al Ca grown in a lathhouse environment at the Tucson Plant Materials Center 32

8. Effect of GA on water requirement values of alfalfa based on yield components and total forage for five harvests in a controlled environment 35

viii

ix

LIST OF TABLES - -C 0111 inu ed

Table Page

9. Effect of GA on dry weight of stein-petiole, leaflet, and total forcige production of alfalfa for five harvests in a controlled environment .... 36

10. Effect of GA on transpired water, 1 eaf].et to stem-petio 1 e .1"atios , and height of alfalfa for five harvests in a controlled environment 37

11. Effect of GA on the anatomy of alfalfa stems 4-3

12. Analyses of variance for the transpiration data, water per cm^ per hr, measured on Mesa-Sirsa alfalfa in a program controlled environment 50

13 • Effect of GA on several, characteristics measured on two clones of Mesa-Sirsa alfalfa grown in a program controlled environment 51

1'i. Summary of the analyses of variance for testing the effect of GA on various characteristics measured on Clone 2 of Mesa-Sirsa alfalfa in a controlled environment 58

15. Effect of GA on carbohydrate fractions and protein content of roots from Clone 2 of Mesa-Sirsa alfalfa grown in a controlled environment 6l

16. Effect of GA on yield of Mesa-Sirsa alfalfa grown under field conditions at Tucson, Arizona for two harvcst periods in .1970 6'±

17* Effect of GA on yield components and leaflet to s t em-pe t.io.l e ratios of Mesa-Sirsa a.l fal. fa grown under field conditions 65

l8. Effect o C GA on the anatomy of Mesa-Sirsa alfalfa leaves and stems 68

X

LIST OF TABLES--Continued

Table Page

19* Effect of GA on chlorophyll content of primary and secondary leaves of Mesa-Sirsa alfalfa 77

20. Effect of GA on carbohydrate fractions and protein content of roots of field grown Mesa-Sirsa alfalfa 80

ABSTRACT

Plants of Medic.ago sativa L. cultivar 'Mesa-Sirsa'

were grown in lathhouse, greenhouse, growth chamber, and

field environments and were used to evaluate the effect of

various antitranspirant and growth regulator chemicals on

water-use efficiency and on physiological, morphological,

and anatomical characteristics. Growth regulators used

were gibberellic acid (GA), indoleacetic acid (IAA) , and

( 2-chloroethyl) trime thy .1. ammonium chloride (CCC). Anti­

transpirant chemicals were phenylinercuric acetate (PMA)

and dodecenylsuccinic acid (DSA). Gibberellic. acid

significantly lowered the water-use efficiency of alfalfa

in all environments. Indoleacetic acid, CCC, PMA, and DSA

did not reduce the amount of water required to produce a

unit of dry forage. The antitranspirant chemicals did not

reduce the amount of water transpired, and further, DSA

caused plant damage when used as a foliar spray.

Gibberellic acid significantly increased the amount

of stem tissue produced in all environments which resulted

in more total dry-forage production. No effect weis found

on the amount of leaflet tissue produced except in experi­

ments where GA stimulated secondary branching.

Flowering was promoted by GA apjilication and

morphological changes were evident in the flowering raceme.

x.i

Pith cell length, leaf thickncss, internode length, stem

length, and rate of stem elongation were increased by GA.

The effect of GA on transpiration was influenced

by soil moisture level. Transpiration rates were higher

on plants treated with GA when 30 to 55% of the available

soil moisture was depleted.

Photosynthesis, respiration, and specific leaf

weight (SLW) were significantly decreased by GA treatment.

— 2 —1 Photosynthetic rates (mg CO^ dm " hr ) were always less

on plants treated with GA. However, when the data were

expressed on a unit of dry leaflet tissue, GA did not

consistently reduce photosynthesis. The time of measure­

ment after treatment and the ago of the plants influenced

photosynthesis aiad respiration rates. The SLW was lower

on plants treated with GA.

Gibbcrellic acid caused a reduction in the chloro­

phyll 13 content of alfalfa leaves. Extreme chlorosis was

noted under field conditions.

Roots from alfalfa plants sprayed with GA had lower

percentages of free and acid-hydrolyzable ca.rbohydratcs

when grown under growth chamber and field conditions.

Sucrose was the only sugar found in the 00% ethanol

extract. Percentage of crude protein was higher in the

roots of plants treated with GA.

INTRODUCTION

Alfalfa (Medicuft'o satri.va L.) .is the most .important

irrigated foi~age crop grown in Arizona. Alfalfa has the

highest consumptive use of water of all the major crops

grown in Arizona (19). Consumptive use of water by

alfalfa is high because of a long growing season and high

yields. Also, in the semiarid climate evaporation is high.

Alfalfa is a high quality feed for livestock con­

sumption. However, if water becomes limited alfalfa

production may be shifted to an alternative forage which

has a lower water consumption. Therefore, it would be

highly desirable to find an alfaIf a cultivar that was more

efficient in water utilization. If water consumption by

present high yielding cu.ltivars could be reduced without

reducing yields, or yields could be increased without in­

creasing water consumption, then the production of alfalfa

in areas with limited water would bo greatly enhanced. The

above factors would also apply to other crops in either

natural rainfall or irrigated areas. The terms water

requirement and water-use efficiency are used inter­

changeably .

This study was initiated with the following

ob j oct .i.vcs :

1

To determine the effect of antitranspirant and..

growth regulator chemicals on water-use efficienc

of alfalfa.

To determine the effect of antitranspirant and

growth regulator chemicals on morphological,

physiological., and anatomical characteristics of

alfalfa.

REVIEW OF LITERATURE

Water Requirement of Alfalfa

There are approximately 80,000 hectares of alfalfa

in Arizona. Alfalfa in the Salt River Project utilizes

188.7 cin ( 7 'i. 3 inches) of water per year (19) - Alfalfa

production creates a large demand for water in an area

where water is becoming limited for agricultural purposes.

The water requirement of alfalfa varies with

different environmental conditions (9)* The literature on

water requirement in both greenhouse and field conditions

has been reviewed extensively (l, 11, 32). The water re­

quirement for alfalfa varied from 657 to l,068over a 7-year

period ('±1) . Al-ICawaz (l) reported significant differences

in the water-use efficiency of alfalfa grown with varying

moisture regimes. Genotypes within a cultivar exhibited

as much variation as differences among cultivars (ll).

Minimum water-use efficiency values were associated with

maximum production of dry forage in most species.

Kelley (33) listed several factors which .influence

the water requirement of plants such as soil moisture,

soil type, fertility, and various climatic conditions.

Other factors such as disease, salinity, chemical agents,

and stage of growth also influence water utilization. Thorn

3

cuid Iloltz (6:i.) concluded that any condition which disturbed

the normal processes of the plant affected the water

requirement.

Effect o f Cheriiic- a Is on Water-Use Efficiency

Several chemicals applied to various species have

affected water utilization (59)• Plants of sorghum

(Sorghum vulgare Pers . ) treated with dalapon (2,2-dich.loro-

propionic acid) had a higlier water requirement in l6 days

(3376) compared to the nontreated (65^) plants (68). Coiner

(.12) found no effect of pyrazon (5-ami]io-'l-chloro-2-phenyl-

3(2H)-pyridazinone) on the water-use efficiency of sugar-

beet (Bet a vulgaris L.) when grown in a greenhouse environ­

ment. Olsen et al. (48) used hexodecanol and octodecanol

on corn (Zea mays L.) and found no reduction in water

utilization; however, production was lower at high rates

of application.

Brcngle (8) found no effect of phenylmercuric

acetate (PMA) on water use by wheat (Tr.itic.u111 nestivum L. ) ,

but damage to the ears and a slightly lower yield were

observed. Swoet orange (Citrus sinensIs Blanco) treated

with latex or silicone coatings exhibited a lower water

requirement (39)* These investigators reported leaf burn

and morplio.log.ical changes in the treated plants.

Plant and llalevy (53) used two growth retarding

chemicals on wheat and found no effect 011 the water-use

efficicncy. Other experiments with o. growth retardant have

shown an interaction between chemical treatment and

moisture regime of the soil (^k).

Ti'ansplratlori Rates of Alfalfa

Very little information is available in the

literature on the transpiration rates of alfalfa expressed

on a leaf area basis. Most of the literature on water use

by alfalfa deals primarily with total consumptive use or

the evapotrans pir at ion rates per day.

Ehrler (.17) reported differences in the amount of

water absorbed by cultivars . 'Lahontan 1 utilized less

water than 'Moapa' in two experiments; however, no values

were reported for either cultivar. lie further reported as

the saturation deficit varied from 0 to 50 nib, transpiration

_ 2 -1 increased from 0 to 300 g m hr . He reported a decrease

in transpiration rates as root temperatures were lowered.

Al-Kawaz (.1) found rates among genotypes of 'Mesa-

— 2 —1 Sirsa 1 to vary from 1.00 to 'l.l4 g H^O dm hr However,

these results should be viewed with caution as they are

single plant observations in an unreplicated experiment.

Al-Kawaz showed a decrease in transpiration rates as the

plants .reached maturity when averaged over all genotypes

within a growth stage.

6

Effect of Chemicals on Transpiration

No data are available on the transpiration rates of

alfalfa per unit of leaf tissue as affected by chemical

treatment. Hales (26) showed a significant reduction .in

the total amount of water used by alfalfa when treated with

various antitranspirants; however, these data gave no

indication of the amount transpired per unit area of leaf.

Some chemical treatments caused foliar damage and reduced

growth which may have reduced the total amount of water

utilized.

Zelitch and Waggoner (7'0 studied the effect of

various chemicals on the transpiration rate of various

species. They found PMA and other compounds reduced

transpiration in some species. For example, PMA reduced

transpiration of tobacco (N i c o (. :i. an a t abacuin L.) in both

leaf disks and intact leaves (73, 7^). Photosynthetic and

the relative growth rates varied with different environ­

ments. Shimshi (5$) confirmed the data of Zelitch and

Waggoner where he noted a reduction in transpiration of

tobacco and Helianthus animus L. when grown in an open

environment. Turner and Waggoner (Gh) reported a 10%

reduction in water use by Plnns resinosa Ait. when sprayed

with PMA under field conditions.

Friesen and Dew (ii2) observed a reduction in

transpiration of a weed species when treated with two

herbicides . The slower ,acting herbicide did not reduce

transpiration as rapidly as the other herbicide. Todd and

Propst (62) noted a reduction in transpiration of leaves

Asrhich were subjected to treatment with o'/->one and ozonated

hexene . Atrazine [ 2-ch.lor o-4- ( ethyl amino ) -6 - ( is opropyl -

amino)-S-trianine] caused a reduction in transpiration of

corn and soybeans (Glyc ine max Morrill) by h-h and 67%,

respectively (59)-

Livne and Vaadia (35) found a differential response

in barley (Hordeum vulgare L.) depending upon the type of

chemical used. An increase in transpiration occurred with

kinetin and gibberellic acid (GA), no response was noted

when adenine and indoleacetic acid (1AA) were used, and a

reduction in the rate occurred when actinomyc.in D and

puromycin were applied to the .leaves. Luke and Freeman

(37) observed no effect on transpiration of excised oat

(Avena satlva L.) leaves when treated with GA; however, a

stimulation was noted when cytokinins were applied. Later,

these researchers were unable to show an effect of

cytokinin on dicotyledonous species (38). Nieinan and

Bernstein (46) reported tin increase in transpiration of

bean leaves treated with GA.

The use of long ch;iin alcohols 011 corn and barley

reduced yield more than transpiration ('l7, 48). Gale,

Roberts, and llagctn (24) reviewed the literature on the use

of alcohols as ;intitraiispirajits and concluded, on the basis

of 17 repoi'ts in which all but one found a reduction in

8

growth, that these materials were unsuitable as effective

antitranspirants. These alcohols offer more resistance to

CO^ exchange than to water movement.

Photosynthetic Rates of Alfalfa

The photosynthetic rate of alfalfa varies with the

age of leaves, environment, and the leaf area index (LAI)

(23, 69)• El-Tabbakh (l8) reported no differences between

field and greenhouse plants in rate of assimilation.

He reported a wide range in rates among cultivars , 23 to 5^

— 2 —1 mg COr> dm hr ' , and Mesa-Sirsa had an average of 43 mg

— 2 -1 COg dm hr when measured at 32 C.

Potassium nutrition has been shown to be important

in the photosynthetic process (13)• As potassium increased

from 0 to 5%, an increase in photosynthetic rate was found

— 2 — 1 (8 to 20 mg dm hr "). Stomatal closure may have been

responsible for the low rates at low potassium levels.

Fuess and Tesar (23) found 3-week-old leaves were

less than one-seventh as active as 5-day-old leaves in

oxygen evolution. These investigators also found an

increase in LAI as the plant reached the bud stage of

development. Wilfong, Disown, and Blaser (69) reported

that photosynthesis .increased as the LAI increased to about

4 and then the rates leveled off. Maximum rates occurred

when 915% of the incident light was intercepted.

9

Pearce, Brown, and Blaser (5l) found a difference

in pliotosyntlietic rates between plants grown in the field

and in growth chambers. Young leaves in the field had a

- 2 - 1 rate of 52 compared to 35 mg C0^ dm hr for the same

age leaves in growth chamber environments. However, when

the rates were expressed on a dry weight basis no difference

existed. This difference in results can be accounted for

2 by the specific leaf weight (SLW, mg/cm ). The SLW was

larger in the field than in the growth chamber. Specific

leaf weight increased as the leaves matured (5l)« Pearce

et al. (52) showed a positive correlation between SLW and

pliotosyntlietic rates. Their data showed that the photo-

-2 -1 synthetic .rate varied from 20 to 50 mg CO dm hr ' as the dt

SLW changed from 1.9 to 5»3*

Al-Kawaz (l) reported differences among genotypes

of Mesa-Sirsa in both photosynthesis and respiration. The

rates of both physiological processes decreased with

maturity.

Effect of Chemicals on Photosyivthes:i.s and Res p i rat1on

G.ibberel lie acid has increased the respiration of

hypocotyls and cotyledons in Cucunils sat.i.vus L. and

decreased the rate in the radicle. Also, an increase in

catalase activity was fouvicl (27). Troltarne and Stoddart

(63) found an inci-oa.se in the activity of ribulose-1 ,5-

diphospliate carboxylase i.11 Trifol.j um pr a tense L. They

10

contributed the increase to an overall effect in promoting

protein synthesis rather than a specific action on the

enzyme. These researchers suggested GA may be part of a

system mediating the pbytocliromc system.

Other investigators (25, 'l-2) found no effect on

1 'i manometric measurements of leaf disk and ' CO^ uptake on

leaves treated with GA. However, Coulombe and Paquin (15)

noted an .increase in respiration and photosynthesis 1 to 2

hours after GA application and maximum effect was noted

5 to 6 hr after treatment. Alvim (2) reported an increase

in net assimilation rates (NAR) of beans (Phas eolus vulgaris

L.) when treated with GA. Ousheva, Popov, and Manolova

((l9) found an increased accumulation of biomass of Ch lor el let

when GA was applied to the media.

Photosynthesis was inhibited when two herbicides

were applied to Scenedesnns sp. The rates were mediated

by light intensity (65) • Todd and Pr*opst (62) found a

reduction in photosynthesis when ozone was applied to leaf

tissues.

Humphries, Weibank, and Witts (29) found a reduc­

tion in NAR of wheat when treated with CCC [(2-chloroethyl)

t r i m e t li y 1 a mm o n i u 111 chloride.]. Plienylmercuric acetate has

been shown to cause ci reduction in photosynthetic rates of

different plant species (57, 59)•

Effect of Chemicals on Alfalfa

Massengale and Medlar (4o) found that TIBA (2,3,5-

triiodobenzoic acid) and 2,4-,5-T ( 2 , k , 5-triclilorophenoxy-

acetic acid) increased stem elongation and IAA caused a

reduction in stem length. No effect on the number of nodes

per stem was found; however, leaf morphology showed some

variation from the nontreated plants. Yeh and Bingham (70)

showed that IAA reduced stem elongation of two genotypes.

These researchers found a clonal interaction in the number

of racemes and florets , and stein height when treated with

GA and TIBA. Other investigators (.10, 21, 71) showed an

increase in stem height in response to GA application.

Corns (I'l) found no response of 'Gi-'imni1 alfalfa to

GA treatment. However, Finn and Nielsen (2l) reported a

.reduction in root growth and an increase in total protein

even though a decrease in percentage protein on a dry

weight basis was found. Carlson, Sprague, and Washko (10)

found a reduction in total carbohydrates when alfalfa was

treated with GA. Yeh (7-1) reported that carbohydrate

levels varied in response to GA application. lie found an

increase in plants grown in the field and a decrease in

greenhouse grown plants.

MATERIALS AND METHODS

Plants of Mesa-Sirsa were used to investigate the

effects of growth regulator and antitranspirant chemicals

on water-use efficiency, transpiration rates, photo­

synthesis, and anatomical features of alfalfa. Studies

were conducted in greenhouse, latlihouse, and gi-owth chamber

environments.

The method of plant cvi.lture was basically the same

in all experiments. Plants were grown in plastic pots with

a hole 1.2 cm in diameter at the base of each pot to alloAf

for drainage during plant establishment. Pea gravel was

added to cover the bottom and a known weight of a mixture

of desert Mohave clay loam soil (Typic Ilap.l argid) , peatmoss,

and organic fertilizer was added to each pot. The ratio of

constituents in the soil mixture was 3 parts soil , 1 part

peatmoss, with .100 g of organic fertilizer (milorganite)

and 3 g of sulfur per 20 kg.

In all experiments there was one plant per pot

which was cut 7 cm above the soil surface at the beginning

of each trial. During plant establishment forage above

the 7 cm level was removed at the 1/10 bloom stage.

Styrofoam was added to the surPace of each pot to reduce

evaporation (.13.) and the pots were watered to bring the

soil to field capacity (27%) at the start of each trial.

13

The experimental design wcis a randomized complete block in

all experiments. Each pot was weighed daily and re-

watered when 55/° of the available: soil moisture had been

utilized. The plants wore harvested at 1/10 bloom and in

each experiment the amoimt of dry forage (80 C for 2k hr)

was divided by the amount of water utilized to detex-mine

the water requirement. Leaflet to stem-petiole ratios were

calculated in all experiments. Height of steins and number

of nodes per stem were determined prior to harvest. Per­

centage N was measured by the micro-lcjeldahl method (3) and

multiplied by 6.25 to estimate crude protein percentage.

Lath and Greenhouse Experiments

Rates and methods of application of chemicals used

in a greenhouse environment are shown in Table 1. The

abbreviation following cach chemical name will be used in

future discussion. All plants received the first chemical

application when regrowth Wcis k cm tall. The second

application was applied k days later. Plants used in this

study were 6 months old and the experiment was conducted in

January, 1969- Average maximum and minimum temperatures

were 3-1 and .1.8 C, respectively. Average relative humidity

was 50/n. Six treatments represented once in each of eight

blocks were us o< 1.

Inform .it ion presented in Table 2 gives the rates

and chemicals used in a lathhouse environment at the Tucson

Table 1. Chemicals, rates, and methods of application used on seeded plants of Mesa-Sirsa alfalfa in a controlled greenhouse environment.

Application rates per plant

Methods of Chemical* application Concentration Low High

Gibberellic acid (GA) f o 1 i ar spray 100 mg/lit er 1 ml 2 ml

Indoleacetic acid (IAA) foliar spray 100 mg/liter 1 ml 2 ml

Phenylmercuric acetate (PMA) foliar spray 1 X H

O 1

1 ml 2 ml

Dodecenylsuccinic acid (DSA) foliar spray 5 X 10 5 M 1 ml 2 ml

(2-chloro ethyl) trimetliylammonium chloride (CCC) soil drench 223 mg/100 ml 100 ml 200 ml

Water foliar spray 0 1 ml 2 ml

*A11 chemicals were used in an aqueous solution.

Table 2. Chemicals and rates used on seeded plants of Mesa-Sirsa alfalfa in a lathhouse environment.

Rates+

Chemical 1 2 3 k

GA* 100 mg/liter 200 mg/liter 300 mg/liter zi00 mg/liter

P>1A* 1 x 10~3 M -4

1 x 3.0 M 1 x 10"5 M 1 X 10~6 M

ccc* 2 M 1 M 1 x i

10~2 M 1 X H

O 1 I-F

-

2

Water plus Tween 20 0 . 2%

W at er 0

*0.2% Tween 20 added to each aqueous solution of chemicals.

+1 ml of each solution was applied as a foliar spray.

H

16

PJ. ant Materials Center in June, 19 6 9 • The numb er of

chemicals was reduced since there was either no effect or

plant damage noted from the chemicals described in Table 1.

The GA, CCC , and control treatments were applied on June

23, 19^9 and the PiMA was applied on June 30, 19^9 • The

plants in this study were 3 months old. Average maximum

and minimum temperatures were 'll and 19 C, respectively.

Average relative humidity was 52%. Fourteen treatments

represented once in each of I'l blocks were used.

Growth Chamber Experiments

Several growth chamber experiments were conducted

to elucidate the significant effect of GA on water utiliza­

tion of alfalfa. One of the first objectives was to

determine if a carryover .response could be measured on

subsequent regrowth. Seeded plants of Mesa-Sirs a were

grown for five consecutive harvests. Two groups, each with

12 plants, were grown in a controlled environmental chamber.

The day and night temperatures were 26 + 1 and l6 +_ 1 C,

respectively, during the 16 hr photoperiod. Light

intensity was 185•8 lux at plant height. The first growth

period was to adjust the piants to the different conditions

of the growth chamber-. During the second growth period,

Group 1 was sprayed with L ml of GA (150 nig/liter) when

regrowth was approximately 3 cm in height. A second

application was applied 'i days later. Group 2 was treated

in the same manner during the fourth growth period. Mar-

vests 3 and 5 were used to evaluate carryover effects.

Height measurements were made at 2-day intervals during

each treated harvest. Stem tissue was collected from the

second harvest to measure the effect of CIA on the anatomy

of alfalfa stems. The samples (0.5 cm) were collected from

an area 1 cm above four consecutive nodes located on the

basal portion of the tallest stem of each plant. Cross

sectional and longitudinal sections were stained with

safranine and fast green according to techniques described

by Johansen (31)* Cells were measured using a calibrated

ocular micrometer.

Transpiration

To measure the effect of GA on transpiration,

propagules of two Mesa-Sirsa clones were established.

These clones were selected for vigorous growth when

spaced planted in the field. Ten propagules of each clone

were grown in the same environmental conditions previously

described. Three con.secut.ive harvests were taken.

The first harvest was used for plant adjustment to

the environment and help in establishing uniformity among

propagules. Two days prior to the second harvest, half of

the propagules of each clone were sprayed with 3 nil of GA

(.150 mg/liter) to wet all leaC foliage. The light period

was from 0700 to 2300 hr . The spraying was in a d e at 2300

18

hr and a3.1 plants were watered to field capacity. The

plants were weighed at periodic intervals to determine the

amount of water loss during the next light period and

rewatered when 55% of the available moisture had been

utilized. Rewatering was at 1700 hr. This procedure was

repeated for 2 consecutive days.

On the third day after the initial spraying, photo-

synthetic measurements were made in an open system using a

Beckman 215 Infrared Gas Analyzer. Measurement of the

entire plant was accomplished by using the plexiglass

chamber shown in Fig. 1. Two air intake-holes were located

near the base and the air was drawn out through a hole near

the center-. Leaf eirea and specific leaf weight (SLW) were

measured by Xeroxing all the leaves on two randomly

selected steins from each plant and determining cm area to

weight ratio. Photosynthesis was expressed as the mg of

2 C0o per dm per hr and also on a dry leaf weight basis C*t

(80 C for 24 hr). Transpiration rates were expressed as

the mg of wat er loss per unit of leaf area per hour.

Photosynthesis and transpiration of the I.eaves below the

7 cm cutoff level were accounted for in the data.

Treatment of the propagules during the third

reg.rowth period was the same .as in the second period.

Propagules wore 5 months old at the beginning of this

experiment.

Fig. 1. Plexiglass chcimber used to measure the photo-synthetic rates of entire alfalfa plants. Note the air intake near the base and exit near the top. The temperature probe is shown in the middle of the chamber.

20

Photosynthesis and Respiration

In the previous experiments, clones reacted differ­

ently to GA treatment. To obtain additional information on

photosynthesis and respiration of GA treated plants, a

study was conducted in Avhich 2h propagules of Clone 2 were

used. The environmental conditions of tb" growth chamber

were the same as those described. The plants were grown

for two consecutive harvests.

The first harvest was used for the plants to adjust

to the environment. During the second growth period half

the plants were sprayed with 1 ml of GA (150 mg/liter) five

days after cutoff when regrowth had begun. The plants were

sprayed at 2300 lir. The treated plants were sprayed again

h. and 8 days later with the same concentration but with a

volume of 1 and 3 nil, respectively, to wet the leaves.

Photosynthesis and respiration data were collected

on four plants of the sprayed and control plants three

times at 4-day intervals beginning the day following the

.initial spray treatment .

All plants were watered to field capacity at 2300

hr of the day preceding measurement.

F i e 1 d E.vp o r ini ent

The field research for this study was conducted at

the University of Arizona Campbell Avenue Farm, Tucson,

Arizona. In October of .1 969 , one border was seeded Av.ith

21

certified Mesa-Sir,Sci alfalfa at 22. k kg/ha. The border was

irrigated to insure sufficient moisture for germination and

einer gene e .

The border was 7-92 x 30 . ;l8 m and it was divided

into six plots (3.66 x 8.53 m)• The plants were allowed to

establish during the winter and the first forage was

removed on April 10, 1970. During the establishment

period, irrigation was managed to prevent water stress.

The border was irrigated during the period of study at the

beginning of each regrowth period . Sufficient water was

applied to fill the entire root zone to field capacity.

No treatments were applied to the regrowth which

developed during the period between April 10 and 25 when

the forage was again removed.

The treatments on the first harvest for data

collection were made on May 15, l6, and 23• Gibberellic

acid was applied with a pressure sprayer at the rate of

2 liters (150 mg/liter) per plot. Three plots wer e sprayed

and three served as the control plots. Forage from a

2 .19 '» and soil moisture samples to a depth of 1.22 m were

taken at 'i-day intervals beginning on May .1.6 and lasting

through May 28.

Ten stems were selected at random from each plot

on May 28 and were used to determine height and internode

length. Stem and Jeaf samples were collected from the

fourth and seventh node and internode from the base to

22

measure the effect of GA on anatomical characteristics.

Also, a leaf was collected fi'om the stem apex. The center

leaflet of each leaf was used in all measurements.

During the second harvest the control plots were

divided in half in order to evaluate the regrowth on the

treated plots of the first harvest. Gibberellic acid

application was the same as that previously described

except the volume was reduced to 1 liter per plot.

Gibberellic acid was applied at k-day intervals beginning

on June 8 and lasting until June l6.

Forage and moisture samples were taken only on

June 21. Beginning on June 9, and at two intervals there­

after until June 21, one stem was selected at random from

each plot, placed in a 125 ml polyethylene bottle filled

with water, and used to determine photosynthetic and

respiration rates. The stems were cut .in the field at 2000,

held in a dark room at 25 C, and measurements were made the

following day between 0800 and 1000 hi'. A plexiglass

chamber shown in fig- 2 was used in the system previously

described for the growth chamber studies.

On June 21, ten stems were selected at random from

each plot for height and internode length measurements.

Additional forage was collected for chlorophyll determina­

tions as described by Seitz (5^0 • Chlorophyll determina­

tions were made on main and secondary leaves separately.

23

Fig. 2. Plexiglass chamber used to measure photosynthetic and respiration rates of plants grown in the field.

2'l

2 Roots from a .19 in ai~ea in all plots were dug at

the end of each regrowth period. The roots were immedi­

ately washed, placed on dry ice, and stored at 0 C until

they were dried in a freeze-dryer. The samples were ground

to pass through a ll0 mesh screen and carbohydrate determina­

tions were made using extraction techniques described by

Dobrenz (l6) and color.imetric determinations described by

Yemon and Willis (72). Qualitative determinations on the

free sugar extract were made by thin layer chromotography

methods described by Stahl (6o). The samples were spotted

on plates coated with silica gel G and impregnated with

0.02 M sodium acetate. The plates were developed with

solvent No. 6 described by Stahl (60). The sugars were

detected with aniline-diphenylamine spray reagent (60).

RESULTS AND DISCUSSION

IJathhouse and Grcnnliouse Experiments

Gibbe.roI.lic acid significantly affected the wate.r-

us e efficiency of alfalfa (Table 3) • Tlie lowest water

requireinent va.Iue was found with the high rate of CIA

application. Plants treated with GA were lower (P - .05)

in water requirement than all other plants treated with

antitranspirnnt and growth regulator chemicals when

averaged over both rates. No effect on water requirement

was found when the plants treated with PMA, IAA, CCC, and

DSA were compared to the control plants,

Plants treated with GA produced the most stein-

petiole tissue (Table k) . A reduction .in stem-petiole

weights were found with the high rate of chemical, applica­

tion. A significant chemical by rate interaction showed

that GA treated plants produced more stem-petiole tissue

at the high rate of application.

None of the chemicals influenced the amount of

leaflet tissue produced when averaged over both rates

(Table h) . However, a chemica l, by rate interaction, was

evident. Maximum and minimum leaflet production was noted

OJI plants treated with CCC at the low and high .rates,

respectiv e1y.

26

Table 3* Effect of various chemicals on water-use effi­ciency of Mesa-Sirsa alfalfa grown in a green­house environment.

Water-use efficiency

Chemic al

Rate CCC GA IAA DSA PMA Control Mean

Low 912* 893 939 ab ab a

High 1072 764 1071 a b • a

992 a 829 b 1005 a

915 9'I8 104-5 9Z12 ab a a

1043 986 1010 991 a a a

979 a 967 a 1027 a

*Values within rates and means followed by the same letter are not significantly different at the .05 level according to Duncan's Multiple Range Test.

27

Tabic k. Effect of various chemicals on. stoni-petio.le and leaflet weight of Mesa-Sirsa alfalfa grown in a greenhouse environment.

Chemical

Rat e CCC GA IAA DS A PMA Control Mean

Stem-petiole weight , ,c r

Low .81 * ab

.75 .80 ab ab

• 70 b

• 72 b

.62 b

• 73 a

High • 51 b

1.00 .52 a b

•51 b

.60 b

.60 b

.62 b

M e an .66 b

.87 .66 a b

. 6l b

. 66 b

• 6l b

Leaflet weight, g

Low • 7'i a

•52 .73 ab a

. 6 6 ab

• 57 ab

.60 ab

.64 a

Hi gh .'19 b

•71 .51 a ab

• 51 ab

• 56 ab

.56 ab

.56 a

.62 a .62 a .62 a .58 a .56 a .58 a

^Values within rates and means followed by the same letter are not significantly different at the .05 level according to Duncan's Multiple Range Test.

28

Total forage production was lower (P = .05) when

high rates of these chemicals were applied (Table 5)•

Maximum forage production occurred on GA treated plants at

the high rate of chemical application.

Leaflet to stem-petiole ratios were significantly

affected (P = .05) by chemical application (Table 5).

Alfalfa plants treated with GA had a lower leaflet to

stem-petiole ratio than any of the other chemical treat­

ments or the control. The lowest ratio was found with

plants treated at the high rate of GA.

All chcmicals tested did not influence the number

of nodes per stem, number of stems per plant, or transpired

water. Plants sprayed wit h GA had longer stems (35 cm)

compared to untreated plants (23*3 cm).

Water-use efficiency was negatively correlated

(P = .01) with total forage, leaflet weight, and stem-

petiole weights. The r values were -0.5$, -0.'l2, and

-0.68, respectively.

The results of this study indicated that GA

affected the .metabolism of alfalfa which was manifested in

an increased plant height and total forage. Increased

forage production was due primarily to an increase in the

amount of stem-petiole tissue produced. Transpired water

was not affected by chemical treatments; therefore, water

requirement was influenced by the significant effect of

these chcmicals on dry forage production.

Table 5- Effect of various chemicals on total forage per plant and leaflet to stem-petiole ratio of Mesa-Sirsa alfalfa grown in a greenhouse environment.

Rat e

Chemic al

Rat e CCC GA IAA DSA PMA Control Mean

Total forage.

Low 1.55* 1.27 1.53 1.35 1.29 1.21 1-37 a ab abc ab abc abc be

High . -99 1.72 1.03 1.02 1.15 1.16 1.18 b c a c c be be

1.27 a 1.49 a 1.28 a 1.19 a 1.22 a 1.19 a

Leaflet to stem-petiole ratio

Low- •95 • 7^ •9^ • 93 .81 •99 .89 a abc cd abc abc bed ab •

High • 99 • 70 1.01 1.06 .96 •96 • 95 a ab d ab ab abc abc

• 97 a .72 b • 97 a 1.00 a .88 a . 97 a

*Values -within rates and means followed by the same letter are not significantly different at the .05 level according to Duncan's Multiple Range Test.

30

Jndoleacetic acid had no significant effect on any

of the characteristics measured except a slight increase

was found in stein height. These results arc: not in agree­

ment ivitli those of other investigators ( 'j0 , 70) . The

antitranspirant (DSA) caused severe leaf burn when sprayed

on alfalfa. Since these chemicals proved ineffectual in

the improvement of water use, IAA, and DSA were not used in

subsequent experiments.

Gibberellic acid, PMA, and CCC had significant

effects on water requirement, total forage, and stem-

petiole weight when a1fa1fa plants were grown in an open

environment (Table 6). The lowest water-use efficiency

values were found with plants treated with GA and the

highest values were found on plants treated with PMA and

CCC. The range in values varied from 173^ to 2^0k for GA

and PMA treated plants, respectively. Water-use efficiency

values in the experiment indicated a strong influence of

the environment on this characteristic. Briggs and Shantz

(9) reported differences in water requirement because of

different environmental conditions.

Maximum forage and stein-petiole tissue production

occurred on GA-treated plants. Different rates of GA

influenced the amount of stem-petio1e tissue. Minimum

forage production occurred on plants treated with CCC.

Leaflet to stem-petiole ratios, stem height, and

transpired water values are shown in Table 7* The lowest

31

Table 6. Means of water-use efficicucy, total forage, and stem-petiole tissue as affected by various chemicals applied to Mesa-Sirsa alfalfa grown in a lathhouse environment at the Tucson PI ant M a L er i a 1 s C enter .

Treatment * Water-us e efficiency

Total forage r

Stem-petiole weight, g

3 1738 +

e 1 . 47 a • 87 a

4 1751 e 1.23 ab • 75 ab

2 1915 de 1.19 abc .67 be

1 2000 c de 1.17 abc .63 bed

l4 2075 bede 1.1k abed •59 bed

11 20 76 be de .89 cd •50 cd

12 2l6l abed .83 d .46 d

10 2198 abed •92 bed .47 d

7 2227 abed 1 .06 bed .56 bed

13 2284 a b c d • 9;i bed .50 cd

9 CO

DO

CO

abc • 96 bed .49 cd

8 2358 ab c • 95 bed .50 cd

6 2 417 ab 1 .04 bed .54 cd

5 2504 a .92 bed .46 d

*1, 2, 3, anc! 4, 100, 200, 300, 400 mg GA/liter, respectively; 5, 6, 7, and 8, 10-61 10-5, 10-4, 10-3 M PMA , respectively; 0, 10, 11, and 12, 10-'1, 10~~, 1, 2 M CCC , res pec I: i.ve.l y ; 13, .2% Twet'ji 20; 1 h , Water.

Values within each column followed by the same letter are not: sigaJ. I' i.cantly different at the .05 level according to Duncan's Multiple Range Test.

32

Table 7- Means of leaflet to stem-pet.i ole ratd.os, height, and transpired water as affected by various chemicals applied to Mesa-Sirsa alfalfa grown in a lathhouse onvironmcnt at the Tucson Plant Mat erials C ent er .

Treatment *

Lcaf1et to stem-petiole

rat io Height , cm Trans pired water, g

4 .65 +

c 19-8 a 2111 abc

3 • 72 be 17.7 ab 2432 a

2 .81 abc 16 .5 abc 2 234 abc

11 .82 abc 16.6 abc 1799 cd

12 .88 ab 15-7 be 1644 d

1 • 91 ab 16. 4 abc 2273 ab

8 .92 cib 13.5 c 2171 abc

l4 • 97 a 15-3 be 2316 ab

7 • 97 a 13.9 be 2234 abc

10 1.00 a 13-7 c 1895 bed

13 .1.02 a 13.0 c 2019 abed

9 1 .02 a 13.9 be 2094 abc

6 .1 .04 a 14.8 be 2335 ab

5 3. . 0 4 a 13 .1 c 2093 abc

*1, 2, 3, mid 4 , 100, 200, 300, 400 nig GA/liter, respectively; 5, h , 7, and 8, 10 ~ 6 , 10-5 , 10~/j, 10 - 3 M PMA, respectively; Q, 10, II, and .12, 10-'1 , 3.0-2, i, 2 M CCC, respectively; 13, .2% Tween 20; 14, Water.

*f* Values within each c.o.lunm followed by the same

letter are not significantly different at the .05 1 eve3. acconl.ing to Duncan's Mul.tip.l.e Range Test.

33

leaflet to stem-petiole ratio (0 .6 5) was found on. GA-

treated plants. The highest rate of GA increased p.lant

height over the control plants. Plants treated with dif­

ferent rates of GA did differ in the amount of water

transpired. The least amount of water was transpired by

plants treated with CCC. However, this was because severe

leaf burn was present on plants sprayed with CCC. In the

previous experiment, CCC was applied as a soil drench and

no visible effects were noted. Plants treated with PMA did

vary from the controls in the amount of water transpired.

This may be due to the length of time in which PMA was

applied to the foliage in the experiment. However, these

results are in agrceinent with the greenhouse study.

Water requirement was associated (P - .01) with

leaflet weight, stem-petiole weight, and total forage. The

r values were -0.55, -0 .76 , and -0.73, respectively. The

significant correlations of dry weight of the total forage

and yie.Ld components with water-use efficiency .indicate

that to select efficient genotypes dry weight production

would be a major selection tool.

None of the chemicals had any effect on the number

of nodes pea" stem, number of stems per plant, or leaflet

weight . Flowering was promoted by GA application.

Tlio results of these experiments indicated that GA

was the only chemical, which improved the water requirement

3 4

of alfalfa without any harmful effects. All other

chemicals either had no effect or caused piant damage.

Growth Chamber Experiments

One of the first objectives of the growth chamber

experiments was to evaluate the carryover effects of GA on

subsequent regrowhli. The int er act ion, treatment by

harvest, showed GA lowered (P = .05) the water requirement

values when calculated from stem-petiole and total forage

weights (Table 0). No effect was found on water require­

ment of each group when averaged over five harvest periods

when the values were computed on yield components or total

forage production. The water requirement values based on

leaflet weights were not significantly affected by GA

treatment.

Gibberellic acid had significant effects on yield

components and total forage of alfalfa (Table 9)• A

significant interaction (P = .05) indicated a positive

response in the amount of forage produced by each group of

plants during the harvest in which they received GA

application.

Transpired water, leaflet to stem-petiole ratios,

and stem height were affected by GA application (Table 10).

More water was transpired by the group of" plants which was

treated with GA. During the throe growth periods when

35

Table 8. Effect of GA on water requirement values of alfalfa based on yield components and total forage for five harvests in a controlled environment.

11 ar vest 1 J. c:UJ. U Group 1 2 3 4 5 M e an

S t em — petiol a*

1 + 1298 . + + b 1193 a 1254 a 1409 b 1354 a 1302 a

2 1121 a 1570 b 1248 a 1229 a i46o a 1325 a

Leaflet

1 1844 a 2020 a 180 3 a 2143 a 2188 a 2000 a

2 1676 a 2079 a 1685 a 1902 a 2099 a 1888 a

Total f or ag e

1 757 b 752 a 736 a 846 b 833 a 785 a

2 669 a 858 b 711 a 74O ci 856 a 767 a

*Values between groups within each harvest followed by the same letter arc not significantly different at the .05 level according to Duncan's Multiple Range Test.

Group 1 treated 2nd harvest Group 2 treated 'id' harvest

Each value is a mean of 12 observations.

36

Table 9* Effect of GA on dry weight of stem-petiole, leaflet, and total forage production of alfalfa for five harvests in a controlled environment.

Harvest j.-' .1. an L Gro up 1 2 3 4 5 Mean

Stem-- it e t .i 0.1 e , K *

i+ 2 . 11 ++ a 2.73 b 3. 46 a 3 .48 a 2 .02 a 2.76 a

2 2 .30 a 1 .83 a 3.72 a 4 . 26 b 2 • 05 a 2.83 a

Leaflet ,, g

1 1 .50 a 1.64 b 2.4l a 2 .29 a 1 .27 a 1.82 a

2 1 . 5 3 a 1 -39

Toti

a

-11

2.6L b

forage, ,1

2 • 75 b 1 .41 a 1 .94 a

1 3 . 61 a 4.3 4 b 5-87 a 5 • 77 a 3 .29 a 4.57 a

2 3 .83 a 3-59 a 6.33 a 7 .01 b 3 .46 a 4.85 a

*Values between groups within each harvest followed by the same letter are not significantly different at the .05 level according to Duncan's Multiple Range Test.

Ciroup 1 treated 2nd harvest Group 2 treated 'it!) harvest

-J* "t* ̂Each value is a mean of .12 observations.

37

Table 10. Effect of GA on transpired water, leaflet to stem~pet:i. ole ratios, and height of alfalfa for five harvests in a controlled environment.

PI ant Group

H arvest

1 4 5 M e an

1

2

2663 a

25'±3 a

+ + l'r nn s | >ir ed wa t er . * .0;

3225 b 4228 a 4-601 a 2701 a 3500 a

2852 a 4343 a 5.I.15 b 2936 a 3558 a

1

2

Leaflet to stem-petiole ratio •

.72 b .60 a .71 a .66 a .62 a .66 a

.67 a .76 b .74 a .66 a .70 b .71 a

.1

2

'17.8 a

4-9.0 a

Height , 0.111

53.2 b 51.0 a 59-3 a 57.4 a

43.9 49.3 cl

53.7 a

62.3 a 55-4 a 52.0 a

*Valu.es between groups within each harvest followed by the same letter are not significantly different at the .05 level according to Duncan's Multiple Range Test.

+Group 1 treated 2nd harvest Group 2 treated 4 <>' liar vest

Each value is a mean of 12 observations.

38

neither group was treated no difference in the amount of

transpired water was found.

The effect of GA on the leaflet to stem-petiole

ratios was not consistent. Differences were found for

harvest 1, 2, and 5• Stem height was increased only on

the second harvest by GA.

Gibbcrellic acid did not appear to have any

influence on any variable measured on the plants during the

harvest following GA application except on leaflet weight

in the third harvest and leaflet to stem-petiole ratio in

the fifth harvest.

Gibbcrellic acid hiuJ no significant effect on

protein percentage of either the leaflet o3.~ stem-petiole

tissue. However, total protein, was increased in the

plants treated with GA and followed the same pattern as

the dry weight changes in each harvest.

Tli e .results of th es-e experiments indicated that the

response of alfalfa to GA treatments was similar to numerous

other species. Increased stem and leaf]et dry weights have

been reported with bean (2). Evtushenko (20) reported GA

was not transferred from one stem to another o11. the same

plant and response was limited to growing parts above the

level of application . Add.i. tiomtlly no carryover effect

was found. Lovell and Booth (36) found no change in the

number of nodes per stem in potato (Sol.anuin tuberosum L.) .

The length of individual internodes was affected by

GA (Fig. 3)» Most of the internodes increased in total

length which result,ed in the increased plant height rather

than more nodes per stem. The largest differences in

internode length were found nearest the base of the stem.

This agrees with Bachelard 's (h) results in. Eucalyptus

c am a 1 d u I. o i1 s 1 s Delia. He reported differences in internode

length depending on the location on the stem and time of

GA treatment. Moore (43) found similar results in the

'Alaska' pea.

Stem length was increased by GA treatment (Fig. k).

Larger differences in stem length were found between the

treated and control, plants as the plants reached maturity.

Different rates of stem elongation were found when alfalfa

plants were sprayed with GA (Fig. 5)- Plants treated with

GA showed a faster rate of elongation h- days eifter spray­

ing. Gibberellic acid was applied on the 7th and 1.1th day

following cutoff. The increase .in stein length occurred at

different rates depending on time of measurement and GA

application. Moore (^3) reported there wore periods wrhen

peas were more sensitive to GA application. The response

of alfalfa to GA app ears to be mediated by time.

The data for cell lengths and the number of xylem

cells per vascular bundle are shown in Table 11. in

general, plants treated with GA had longer pith cells.

However, some genotypes did not respond to GA application.

KEY: • A = TREATED A = CONTROL

CI

O o

CD

o 'S3

o

c

1 2 3 4 S 5 7 S S 10

INTERNODE LOCATION

Fig. 3- Effect of GA on length of individual internodes of alfalfa grown in a controlled environment. Internodes are numbered consecutively from cutoff level (one) to stein tip (ten). «£-

6

KEY : © = TRERTED a= CONTROL

DAYS RFTER CUTOFF

F i g . k . Effect of GA oil height of alfalfa steins grown in a control.led environment .

KEY: Q= TREATED a= CONTROL

DAYS RFTER CUTOFF

Fig. 5 . Effoc.t of GA on the rate of stem elongation in alfalfa grown in a controlled environment.

'i3

Table 11. Effect of 01A on the anatomy of alfalfa st ems.

Plant group

Internode Plant group 1 + 2 3 k Mean

Pith cell length, 111111

Treated . 3 8 a* .36 a .29 a .30 a •33 a

Control • 35 a . 30 a •32 a .25 a . 31 a

No. of xylem cells per vascular bundle

Tre exted 23.9 a 22.0 a 20.0 a 17-6 a 20.9 a

Control 22.7 a 21.1 a 19-0 a 16.2 a v.0 CO

*Vc»Iues between groups within each internode followed by the same letter are not significantly different at the .05 level according to Duncan's Multiple Range Test.

Base of plant.

kh

The longest colls were located in the internodes (l and 2,

Tabic 11) nearest the basal end of the stem and decreased

toward the stem apex. The results of cell length and

inter node length suggest, that cell elongation was primarily

responsible for the increased length. Bos track and

Struekmeyer (7) reported that GA application caused cell

elongation and a smaller stem diameter in various plant

species. Earlier, these investigators had reported

similar results for soybeans (6).

Essentially no d i ff e r e n c e s were found in the

number of xylein cells between the treated and control

plants. Morey and Cronshaw (kh) found that GA had no

effect on the amount of secondary xylem in Acer rubrum and

GA stimulated cambial activity in localized regions of the

stem.

Effect of GA on Transpiration

Transpiration rates were not constant for two

clones of alfalfa (Fig. 6). M aximum rates occurred during

the first hour following the dark period. The rates

declined during the day as soil moisture tensions

increased. The plants were rewatered at ]. 700 hr. During

the period between 1700 and 2000 hi- transpiration rates

increased because of the lower soil moisture tensions. A

rapid decline occurred during the period between 2000 and

2300 hr. Differences (P - .05) between the clones occurred

KEY: 0 =CLONE 1 a = CLONE 2

TIME (HOURS3

F ig. 6. Transpiration rates for two clones of Mesa-Sirsa alfalfa grown in a controlled environment.

4-6

at 1100, I'lOO, 1700, and 2300 hr . No differences were

found during the dark period. Transpiration rates between

clones were larger1 when 30-55/" of the available soil

moisture was utilized. Selection of alfalfa genotypes

which have low transpiration rates should be done at low

soil moisture levels. However, the difference found in

these clones may not be sufficient to detect under field

conditions.

The rates reported by Al-Kawaz (1) arc lower than

those of the present study. Ehrler (.17) showed differences

in the amount of water absorbed as the saturation deficit

changed. Therefore, difference between the reported rates

can probably be attributed to the different environments.

Gibberellic acid affected the transpiration rates

of alfalfa (Fig. 7)- I", general, higher rates were found

on plants treated with GA. Higher transpiration rates have

been reported in tomato and beans when treated with GA

(15, '16). However, Luke and Freeman (3$) found no effect

of GA on transpiration in oats (Avena sativa L.) .

Each clone responded differently to GA application

(Fig. 8). Clone 2 showed a larger increase in the trans­

piration rates with GA treatment than did Clone 1. These

experiments were conducted over a 2-day period and Clone 2

responded similarly both days. However, Clone 1 showed a

decrease the first flay and cin increase the second day,

which caused a significant day by clone interaction.

KEY: 0 =CONTROL A = TREATED O C7

O c

c

CM CTJ

o

o o

2330

TIME (HOURS)

Fig. (. Effect of GA on the transpiration rates of Mesa-Sirsa alfalfa grown in a controlled environment.

t

CNTRL-CLN2

TIME (HOURS)

Fig. 8. Effect of GA on the transpiration rates of two clones of Mesa-Sirsa alfalfa grora in a controlled environment.

CO

iL 9

(P = .05). A summary of the analyses of variance for each

time period is shown in Table 12. The experiment was

repeated with the same clones and similar results were

found.

Data for several characteristics, measured on two

clones used in the transpiration experiments, exhibited

variable responses to GA treatment (Table 13)« No differ­

ences were found in several of the characteristics

measured. This was expected because GA was applied only 2

days prior to harvest. However, differences were found in

the photosynthetic rates. A significant clone by treatment

interaction was found. This interaction was evident

whether the data were expressed on a leaf area or leaf

weight basis. The highest rate of photosynthesis was found

in Clone 2 treated with GA. Clone 1 and Clone 2 showed a

decrease and an increase in the photosynthetic rates,

respectively, in response to GA application.

Differences were found in the SLW of the leaves.

The SLW was lower in the treated plants of Clone 1 and no

differences were found in Clone 2.

Effect of GA on Photosynthesis and Respiration

Effects of GA on several parameters measured on

Clone 2 of Mesa-Sirsa alfalfa are shown in Figs. 9 to .1 lL.

Significant differences for all characteristics except SLW

and photosynthesis were found among harvests (Table .1 k) .

Tabl e .1.2. Analyses of variance for the transpiration data, wat or per cm" per hr, measured on Mesa-Sirsa alfal fa in a program controlled environment .

50

Time

Components 0700 OFLOO 1100 I'JOO 1700 2000 2300

Clone ns ns * * * * * ns **

Treatment ns ns ns * * ns ns ns

Clone by treatment ns ns ns ns ns ns ns

Day lis * * ns ** :|; * ns ns

Day by clone ns * ns ns ns ns ns

Day by treatment ns ns ns * ns ns ns

Day by clone by treatment lis ns ns ns ns lis ns

ns Not significant.

* S i gn .1 f i c ant .05 level.

* * Si gni f :i c ant .01 1 evel .

Table 13 • Effect of GA on several characteristics measured on two clones of Mesa-Sirsa alfalfa grown in a program controlled environment.

Clone 1 Clone 2

Characteristics Control Treated Control Treated

Stem-petiole weight, g Leaflet weight, g Total forage, g Transpired water, g Water requirement Leaflet to stem-petiole ratio Per cent moisture Apparent photosynthesis' Apparent photosynthesis + +

Specific leaf weight, mg/cm Leaf nrea. cm-

2 .02 a* 1 .88 a 1 .88 a 1 .76 a 1 .13 a .90 a 1 .02 a .88 a 3 .15 a 2 .78 a 2 .90 a O .64 a

1279 s. 109'5 a 1567 a 1312 a 395 b 384 b 539 a 496 a

.56 a .48 b . 5 'i ab . 50 ab 80 . 1 a 81 . 8 a 80 . 8 a 80 . 1 a 8i . 7 b 79 .1 b 90 . 9 ab 100 . 7 a 35 .1 a 27 .8 b 29 . 7 ab 35 .2 a k • 3 a n J . 6 b 3 • 3 b 3 • 5 b

260 a 253 a 309 a 250 a

*Values within characteristics followed by the same letter are not significantly different at the .05 level according to Duncan's Multiple Range Test. Each value is a mean of five observations.

' mg C0o g hr

mg C0r> dm " hr

Kb f o = i Rc.R i c.0 * = CONTROL

rw p To •—r o

UJ y UJ al

LU O a:

i cr

CONiROL RESTED

10 K

DRYS RFTER CUTOFF DRYS RFTER CUTOFF

Fig. 9* Effect of GA on the percentage moisture and total forage of Mesa-Sirsa alfalfa grown in a controlled environment.

cc

KEY: © = TRERTED * = CONTROL KEY: 0= TRERTED ^ = CQNTRO'

!

C)

LiJ

CE LJ

1 14

DRYS RFTER CUTCF DRYS RFTER CUTOFF

Effect cf G.A on the s'ceni-peticle and leaflet weights of Mesa-Sirsa alfalfa grov.'n in a controlled environment .

i

KEY: e = TRERTED a =CONTROL KEY : o =TREATED A =CONTROL

! i J cr: ZD o i;J

OC L'J ! -o: ~j I

DRY-S RFTER CUTOFF GRYS RFTER CUTOFF

11. Effecl of GA on transpired vrater and water requirement of Mesa-Sirsa alfalfa grovrn in a controlled environment .

\JI

KEY: o = IRERIED ^ = CONTROL KEY: o=TREATED a = CONTROL

C3 00

cb

to

1^ 10

1 14

DRYS fiFTER CUTOFF DRYS RFTER CUTOFF

Effect of GA on leaflet to stem-petiole ratio and specific of Wcsa-Sirsa alfalfa grown in a controlled environment.

leaf weight

KEY: o = TRERTED a = CONTROL KEY: o=TRERTED a = CONTROL

o

cn ai i—i Q_ CO UJ

O

CE C£ i—i Q-CO UJ CC

DRYS RFTER CUTOFF DRYS RFTER CUTOFF

Fig. 13' Effect of GA on respiration rates of Mesa Sirsa alfalfa grown in a controlled environment. Left) mg CC>2 dm~2 hr~l. Right) mg CC>2 g-1 hr~l

ui C7\

KEY: o= TREATED a = CONTROL KEY: o = TRERTED ^ = CONTROL <s

CO

o CJ

CO LU x: o

CM ~ CO I— ^— CO o I— o 32 Q_ o

o

10 14 6

o

CO

o

CO i—i

o CO UJ 31 CO I— >-CO o I— o O 32 Q_

LO

O d

DAYS AFTER CUTOFF , DAYS AFTER CUTOFF

Fig. l4. Effect of GA on photosynthetic rates of Mesa-Sirsa alfalfa grown in a controlled environment. Left) mg CO dm-^ hr~l. Right) rag C0o g~l hr~l.

u Z

VJ1

Table 1 . Summary of the analyses of variance for testing the effect of GA on various characteristics measured on Clone 2 of Mesa-Sirsa alfalfa in a controlled environment.

Characteristic Treatment Harvest Treatment by harvest

Fresh weight, g * * * ns Transpired water, g ns * 11 s Stem-petiole weight, g * *

Leaflet weight, g ns * * ns Total forage, g ns * * 11s V."ater-use efficiency * *

Leaflet to stem-petiole ratio ns * * ns Specific leaf weight, mg/cm^ ns * *

Per cent moisture lis * * ns Respiration, mg CO2 dm-^ hr~l ns * * ns Photosynthesis, mg CO2 dm~2 hr*"l ns ns ns Respiration, mg CO2 g~hr-1 ns * * ns Photosynthesis, mg CO2 hr-1 ns ns ns

ns Not significant.

*•05 level of significance.

**.01 level of significance.

59

The data for total forage, each component of yield, and

transpired water show an approximate linear increase with

maturity. The plants Avere in the bud stage of development

at the final harvest. The plants treated with GA had a

higher percentage moisture and the difference became larger

with .increased plant age (Fig. 9)» Shchulcina (55) found a

lower percentage moisture in the leaves of alfalfa when

treated with GA.

Water requirement values for the treated plants

remained constant over time compared to the increase in

nontreated plants (Fig. 11). The reduction in water

requirement was primarily due to increased stein-petiole

production with no difference in the amount of water

transpired or leaflet production. The leaflet to stem-

petiole ratio of the treated propagules was not signifi­

cantly lower than the control; however, the trend was a

lower ratio in the treated plants.

Differences were found in the SLW between treated

and control plants and the effect became larger with time

(Fig. 12). The SLW was always lower for the treated

plants. Pearce et al . (52) reported that the SLW A^aried

with leaf age and was influenced by light intensity. They

reported SLW values varied from 1.9 to 5«5« The SLW

values of this experiment varied from 2.0 to 2.9.

Respiration differed among harvests whether the

data were expressed on leaf area or leaf weight basis

6o

(Fig. 13). The same gcnural pattern was evident in both

methods of data expression, with the control plants lower

.in the second and thi.rd harvests . Respiration values

- 2 - L varied from 1.0 to 3*1 CO0 din hi" " . These values

C-t

compare to those reported by Al-Kawaz (.!)•

Photosynthctic rates were always lower on plants

treated with GA when the data were expressed on a unit of

leaf area (Fig. 14) . However, whan the d at a were expressed

on leaf weight the control plants were lower in the second

and third harvests. These results can be attributed to a

decrease in the SLW of the treated plants. The same leaf

area of the treated plants did not weigh the same as that

of nontrcated plants. Therefore, on a weight basis there

were more leaves per gram in the treated piciiits .

Coulombe and Paquin (15) indicated that GA influ­

enced the rate of respiration and photosynthesis of

tomato. They reported the maximum effect resulting from

GA application was 5 hr after treatment and then the rates

began to decline to normal. The measurements on alfalfa

wore in a d e approximately 9 hr after treatment; therefore,

maximum response may not have been found in this study.

The percentages of each carbohydrate fraction and

crude protein of the roots were affected by GA (Table 15) •

The .amount of free sugars and the ncid-hydi'olyzoble frac­

tion were reduced by 1 ') . (t and 3f5.8%, res pec Lively . A

slight, increase was found in pro (. e in con Lent. Love 11 and

6i

Table IfJ. Effect ol" GA on c arbohydrat e fractions and protein content of roots from Clone 2 of Mesa-Sirsa alfalfa grown in a controlled environment .

Carbohydrate fractions, %

Plant 80% O 0/ <i/o Protein g.r 0 u p ethanol H2S04

percentage

Treated 12.0 b:i 17.6 b 9.5 a

Control 1 4. 7 a 28.4 a 9-0 a

% of control. 8.1 .4 62.2 105-9

*Values between treatments for each parameter followed 'by the same letter are not significant at the .05 level according to Duncan's Multiple Range Test.

Yi o o t h (36) f o und a reduction in the amount of ethanol

soluble carbohydrates in potatoes when treated with GA.

The results of this study suggest the increase in

stem-petiole tissue was because the root reserves were not

replenished. Previous work und or irrigated conditions in

Arizona lias shown that starch was the major storage compound

in alfalfa roots (lG). Weaver, Shindy, and Kl;i. ewer (G 7)

showed GA had a sign ificant inf luence on translocation of

organic acids in grapes . Other research ( 4 , 36, >15) has

shown GA influenced the amount of starch stored in various

plant organs. The photosynthetic products were probably

not trnjis.l ocatod to the roots and were used to increase the

dry weight of the stem-petiole tissue in alfalfa.

62

F j. e 1 d Exp ei":i.inon t s

Gibberel.l.ic acid (15G nig/I it er) was sprayed three

times in. the f i.e 'l d at 4 day iiitcrva I s beginning who 11 the

regrovth was approxiniately G cm. Gibbercllic acid signifi-

cantly affccted CI.VA,' forage accumulation under .field condi­

tions (Fig. 15)• Plants which were sprayed with GA

produced more forage (P = .05) than did the control plants.

A rapid increase was noted during the first three harvests.

Significant differences in forage yields at the bud and

early bloom stage of plant development for each harvest

were found for plants sprayed with GA (Table l6). The

treated plants yielded more forage than the nontreated

plants (P = .05)- However, regrowth was reduced by GA

treatment.

The effect of GA on components of yield and leaflet

to stem-petiole ratios was also evaluated under field

conditions (Table 17)• In general, primary and secondary

stem weights were greater on the alfalfa plants sprayed

with GA. The total amount of leaf tissue was not influ­

enced in either harvest by GA treatment. The leaflet to

stem-petiole ratios were lower on the treated plants in

both harvests because of an increase in stem weight. The

treated and nontreated plants in Pig. 16 illustrate the

effects of GA on stem height and secondary branching of

alfalfa plants grown under field conditions. The effect of

63

a = CONTROL KEY s o= TRERTED

DRYS RFTER CUTOFF

Fig. 15. Effect of GA on the yield of Mes^-Sirsn alfalfa crown under field conditions. Data "taken from an o area ot .19 in".

64

Table .1.6. Effect of GA on yield of Mes a-Sirs a alfalfa grown under field conditions at Tucson, Arizona for two harvest periods in 1970.

Dry forage production (g/plot)

Harvest Treated Control Regrowth p c r i o d g S 1st treat, ed

May 28 67-5 a* 56.7 b - -

June 22 78.5 a 50-9 b 33.1 c

* Veil ues followed by the same letter within a harvest period ore not significantly different according to Duncan's Multiple Range Test. Mean based on 3 samples collected from an area of .19 "i" (2 ft 2) .

Table 1 7 - Effect of GA on yield components and leaflet to stem-petiole ratios of Mesa-Sirsa alfalfa grora under field conditions.

Primary Primary- S econdary Secondary Total Total Total Harvest Treatment s t ems leaves s t ems 1 e a v e s s t ems 1 eaves forage

Components viol d rr

Treated 8.78 a* 2 . 20 a 2.05 a 2 . 31 a 10.83 a 4.51 a 15-33 a May 28

Control 6.14 b 2 . 34 a .83 a 1.71 a 6.97 a 4.05 a 11.01 a

Treated 7.07 a 1. 30 a 2 . 40 a 1 .81 a 8.37 a 4.21 a 12.58 a June 22

Control 5.08 b .52 b 2.26 a 1-37 b 5 .60 b 3.63 a 9.23 b

Leaflet to stem-p etio1e ratio

Primary- •Secondary- Total axis axis pi ant

Treated .25 b 1 .14 a .41 b May 28

Control .38 a 2.15 a .58 a

Treated .34 b 1-39 b • 50 b June 22

Control .44 b 2.26 a .65 a

* Values between treatments are not significantly different at Range Test.

for the

each parameter followed by the same letter .05 level according to Duncan's Multiple

H • •u HB

Em

•1

66

Fig. l6. Effect of GA on stem length and secondary branching. l) Control. 2) Treated.

67

GA on these parameters was also evident in the growth

chamber studies.

The amount of water utilized by the treated and

nontreated plants was not influenced by GA application.

Significant differences were found in the amount of water

absorbed at different soil depths over the period of the

study. However, no harvest by soil depth-level interaction

was found. Therefore, the plants which were sprayed with

GA had a lower water requirement based on forage produc­

tion .

Anatomical measurements on stems and leaves of

alfalfa indicated GA influenced these parameters (Table

l8). Data from the field experiments agree with the

growth-chamber experiments. Increases in the parenchyma

cell length were evident in the central pith. Also,

individual internodc lengths were comparable to the previ­

ous results shown in Fig. J. Leaflets were thicker 011

treated plants near the base of the stem. Spraying with

GA affected other anatomical features of alfalfa leaves

(Fig. 17)• Intercellular spaces were more abundant in the

treated leaves. Also, the mesopliyll cells were elongated

and less densely arranged. No effect was evident in the

central midvein. Bostraclc and Struckmeyer (6) reported

more intercellular spaces and a smaller palisade layer in

treated soybean leaves.

68

Tabic l8. Effect of GA on the anatomy of Mesa-Sirsa alfalfa leaves and stems.

Int ernod e from base

Pith c: ell length, mm Leaf th.i ickiiess , 111111 Int ernod e from base Tro a10 d C011.tr 101 Treated Control

k .3 yJ a * .29 a .26 a .21 b

7 .32 a .18 a . 20 a .19 a

ap ex — • l6 a .16 a

* V a I u e s between t r c a t monts v i t h in a n in t e mode location followed by the same letter axe not significantly different at the .05 level according to Duncan's Multiple Range Test.

69

Fig. 1 7 . Effect of GA on alfalfa leaf anatomy. Note the increased width and the arrangement of palisade and spongy mesophyll cells (1250 X). Upper) Treated. Lower) Control.

70

Application of CVA definitely inriuenced raceme and

flower development (Fig. -1.8) , No quantitative measurements

were made, but it was evident by visual observation that GA

promoted eaivl lor I:'Ioweriii.g and elongated the flower racemes.

Also, the sepal s were elongated and were curled ne ar the

apex. iS'o differences were evident in flower color.

The effect of GA on photosynthesis, respiration,

and SLW of alfalfa grown under field corulj Lions was investi­

gated (figs. 19, 20, and 21) . Plants treat0d with GA

exhibited lower va.'Laes in. all parameters over most of the

growing period . Differences (P •- .05) between harvests

were found in all characteristics. The results agree with

those obtained in growth chamber studies. The photo-

synthetic and respiration, rates were very similar between

environments except, for the first harvest in the field

where higher rates were found. The high rates were

probably the result of age of the leaves. Previous

research had j.ndic irt ed a decrease in photosyn the tic rates

as 1 ea ves 111 atur ed ( 5 L ) •

The increase in al 1 parameters at the sixth harvest

is d i fficul t (0 explain . The tec.hui.que used in this study

for pliot os yn the tic measurements allowed use of only the

upper 25 to 28 cm of the stem. At; the time of the sixth

harvest, secondary Loaf development was prominent from the

axillary buds located in the; region used for measurement .

Effect of GA on raceme and sepal elongation, l) Control. 2) Treated.

KEY: o = TREATED * = CONTROL o

to

o

o cn CNJ to i—i

to UJ x I—

o cn

t— O X 0-

o LO

D O

24 12 18 22 16 14

DAYS RFTER CUTOFF

_ n Fig. 1 9 . Effect of OA on photosynthesis (mg COo dm *" lir

of alfalfa grown under field conditions.

K E Y : © = T R E A T E D * = C O N T R O L

Csl -J-

P? -cn

_

D

O T 22 12

i 14

i IB 18

I 20 24

DRYS RFTER CUTOFF

- 2 Fig. 20. Effect of GA on respiration (mg COo dm hr

of a] fa] fa grown under field cond i Lions .

7'J

KEY: o= TREfiTED a= CONTROL

DRYS RFTER CUTOFF

Fig. 21. Effect of GA on .specific .Loaf weight (SLW) alfalfa grown under fic.l d conditions.

of

75

The increased number of young leaves may have accounted

for the increased rates.

Respiration and photosynthesis ineasuremouts compni'e

favorably with values reported in the literature for

alfalfa (.1.8, 51). Previous research lias shown that GA

influenced the NAR of beans and wheat (2, 29) . However, a

species response was noted with a decrea.se in wheat and an

increase in beans. IlaJevy, Monsc.l ise, and I'.l.aut (28)

showed a decrease in the di~y matter content of leaves and

attributed the reduction, to increased translocation. The

lower SLW may have resulted from an increased translocation

in alfalfa.

Chlorosis on the treated plots was evident in the

field (Fig-. 22). Chlorosis .in other species treated with

GA has been reported by several researchers (5, 6, ̂ J2, 66).

Reductions in chlorophyll content were found for mono-

cotyledonous and dieotyj edonous species (66). Larger

differences were evident in the chlorophyll 13 content of

primary leaves with little variation in the secondary

leaves (Table 19)• Ismail, Biggs, and Oberbacher (30)

found that GA delayed ch.l orophyl I degradation in detached

oranges ( C.i. I.rus s [ n ens is Blanco) ; however , a cu.'l tivar

response was evident. Results with other legume species ,

peas and vetch, have shown a decrease .in chlorophyll

content (66). A spectrophoLometric trace of a chlorophyll

Chlorosis and increased height of Mesa-Sirsa alfalfa following GA application under field conditions.

Table 19- Effect of GA on chlorophyll content of primary and secondary leaves of Mesa-Sirsa alfalfa.

Leaf type Treatment

C h1or o phy11 A

mg/g F.W,

Chlorophvll B

mg/g F.lv.

Total Chlorophyll mg/g F . \v.

% Chlorophyll

A'

Primary Treat cd

Control

1.20 a'

1.32 a

, 30 b

.8^1 a

1.50 b

2.16 a

80.0 a

61.1 b

S econdarv Treated

Control

1 .20 a

1 . 32 a

.30 a

. 36 a

1 .50 a

I.56 a

80.0 a

8 k . 6 a

letter ar Multiple

Values between treatments within e not significantly different at Range Test.

each parameter the .05 level

' followed by according to

the same Dune an's

7'6

extract indicated that GA a I' fee ted the transmi t1 anc e of

.light at all wavelengths between 4 00 and 700 ni]i. (Fig. 23).

'Jiie effect o C GA on c arbohydrnt e fractions and

percentage: prott:.i.n of field grown alfalfa rooty v,7ere

measured (Table 20). The results of the field experiments

agree with the growth chamber studies. A reduction in each

carbohydrate fraction was evident in both harvests. The

regrowth of the plants treated with GA in the first harvest

was intermediate in. the acid-hydros,able carbohydrate

fraction at the end of the second regrowth period. These

data indicate that starch storage was delayed only for the

first harvest and GA had ny influence on subsequent trans­

location . Percentage protein showed a slight increase in.

the treated plants.

Qualitative determinations of the free sugar

extract indicated that only sucrose was present (Fig. 2fl) .

Dobrenz (.1.6) indicated that sucro.se wets the major component

of the free sugars; however, glucose and fructose were

present in small quantities. In the present study,

sample preparation and a different c.ultivar may have

accounted for the absence of glucose and fructose. Lee

and Rosa (3'0 found a reduction of starch in tobacco

leaves treated with (iA . Other- researchers ( 3 , 'l 5 ) showed

that starch metabolism was affec.ted and more sucrose was

present in. trc?aled corn leaves (50).

79

100 /-1—' 700

Fig. 23* Effect of GA on per chlorophyll extract

cent transniittance of a of Mesa-Sirsa alfalfa.

Table 20. Effect of GA on carbohydrate fractions and protein content of roots of field grora .Mesa-Sirsa alfalfa.

Carbohydrate fractions, %

Harvest Treatment

U fV / uU/0 ethanol "2-50/,

Protein oercent age

Mav 28

Treated

Control

% of control

9 • !i a:

11.1 a

8^.9

6.0 b

18.9 a

31-7

l^i.5 a

13.9 a

104.8

June 22

Treat ed

Control

Regrowth

9-5 b

13-6 a

13-9 a

k.7 c

11.0 a

7.9 b

10. Li a

9-5 a

9-5 a

0' of control 69 .9 ZLO 109-6

'Values between treatments for each parameter followed by the same letter are not significantly different at the .05 level according to Duncan's Nultiple Range Test.

8i

Fig. 2k. Photograph showing the separation of free sugars by thin layer chromatography. Standards were prepared at a concentration of 2 mg/ml. Maltose was spotted with 10 and 25 [il , and M2 , respectively. Sucrose (S) and fructose (F) were spotted with 25 JJ,1 and the unknown was spotted with 5, 10, 25, and 50 |j,l, la, lb, lc , and Id, respectively.

82

The datct of the field experiment indicate that the

water requirement of alfalfa can be lowered by increasing

dry forage production. Under field conditions, GA

increased the total forage produced when applied as an

aqueous spray. However t root reserves were reduced and

regrowth was lower on the subsequent harvest. Therefore,

the application of GA oil a practical basis is not

jus ti Pied.

SUMMARY

Plants of Mo rile a.tco «ativa L . cultxvur Mcsa-Sirs;i

were used to evaluate the offact of various growth regula­

tor ami ant i trai"1 s p Iran t chemicals on water-UH e efficiency

and o tli or physiological. , morphological, and anatomical

characteristics . Experimen ts wore conducted in greenhouse,

lathhou.se, growth chamber, and field environments.

Water-use efficiency values were significantly

lower when plants were sprayed with an aqueous solution of

GA , in all environments. However, when other growth

regulator or antitranspirant chemicals were applied no

sign.i r.i.caut effec t was found on water requirement in a

greenhouse environment. The water requirement of alfalfa

was increased when plants were sprayed with PMA and CCC in.

a lathhouse environment.

Gibberellie acid increased total forage production

in all. experiments . The increase was primar 11.y due to more

stem-petiole production with essentially no change in

leaflet production. However, in some experiments secondary

brandling was increased by GA and this resulted in an

increased amount of leaflet tissues.

Gibberellic acid proniot ed early flowering and caused

morphological changes in tin.- flowering raceme . An increase

in pi.th cell length, .'leaf thickness, i.nternodo length, stem

A 3

8'i

height, ,'ind .rate of stem elonga tioji \fas found on plants

sprayed with GA .

The effect of GA on l;raTiP|):i rati on was mediated by

soil moisture comii Lions. When, the soil mo is turo content

was nenr fx old capacity GA did not inl'J uence transpirata.on

rates. However, as the soil 1110 is turo-porccnt age decreased

transpiration rates were significantly increased, Two

alfalfa genotypes responded differently to GA treatment.

- 2 - I Photosynthesis (nig C0r dm " ar ' ) was I ower for plants

treated with GA when .grown under growth chamber and field

environments. The S1. V was .lower on p ."1 ants sprayed with GA;

therefore . when the photosynthesis and respi.ral.ion data

were expressed on a leaf weight: basis, variable results

were found depending on the time of measurenient after

cutoff.

Chlorosis was evident on pi an Is sprayed with GA in

field conditions. The yellowing was primarily caused by a

reduction in the chlorophyll 1! content, of primary leaves.

Also, all fractions of the chlorophyll extract were lowered

in both primary and secondary leaves.

Plants treated with GA had a .1 owor percentage of

free and acid-hydro.l. yzable carbohydrates in the roots.

This effect was found in both greenhouse and field environ­

ments. No qual i t at :i. v e effect was found on the free sugars

extracted wj th 8()"'i < - than "> I . Sucrose was the only sugar

I) r e s e 111 in t h e e x L r a c t .

85

Percentage protein of the roots showed a slight

inc.rea.se when the plants were sprayed with GA. However, no

effect on protein percentage was evident in the above

ground forage when the plants were treated with GA.

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