genetic diversity of iranian clones of common reed (phragmites australis) based on morphological...

Genetic Diversity of Iranian Clones of Common Reed (Phragmites australis)Based on Morphological Traits and RAPD Markers

Marjan Diyanat, Ali A. S. Booshehri, Hassan M. Alizadeh, Mohammad R. Naghavi, and Hamid R. Mashhadi*

The genetic diversity of 39 clones of common reed originating from different geographical areas of Iran were evaluatedusing morphological and RAPD analyses. High level of morphological variation was observed among clones. The 16primers used in this study amplified 149 scorable RAPD loci among which 123 were polymorphic (83.1%). A dendrogramwas prepared on the basis of a similarity matrix of RAPD data using the unweighted pair-group method with arithmeticaverages (UPGMA) algorithm and separated the 39 clones into four groups, which mainly were in accordance withgeographical origins. The results of the morphological comparison mostly corresponded with the results of RAPD analysis.It is possible that these variations among clones will affect successful management of common reed using chemical or theother methods of control.Nomenclature: Common reed, Phragmites australis (Cav.) Trin. ex Steud.Key words: Morphological traits, RAPD, diversity, cluster analysis.

Common reed is a perennial grass and is the mostwidespread flowering plant in the world (Clevering andLissner 1999). It is found on every continent except Antarcticaand is common throughout Asia, North America, and Europe(Saltonstall 2002). Common reed enlarges its population byclonal growth through rhizomes and is a typical plural clonalplant species (Dong 1996). It now is found in many provincesin Iran and causes problems in some provinces (e.g., Ardebil).

In the last few years, the field of molecular biology hasprovided new tools for studying population structure. Clonaldiversity and evolutionary processes in wetland species (suchas cattain [Typha] and cordgrass [Spartina]) were studied forthe first time, using allozyme polymorphisms (McNaughton1975; Raybould et al. 1991; Silander 1985). Since the 1980s,new perspectives in how to study evolutionary processes andpopulation dynamics in common reed became available withthe development of molecular markers (de Kroon andvan Groenendael 1997; Jackson et al. 1985). However, ourknowledge of the underlying evolutionary processes indetermining the clonal diversity still is limited.

Significant morphological differences have been found bothamong different populations of common reed and differentclones within the same populations, irrespective of siteconditions (Bjork 1967; Clevering 1999; Clevering et al.2001; Hansen et al. 2007; Pauca-Comanescu et al. 1999;Rolletschek et al. 1999). Part of the clone-specific variabilityin common reed can be attributed to differences inchromosome number. A euploid range of 3x, 4x, 6x, 7x, 8x,10x, 11x, and 12x (with x 5 12) has been found for thisspecies, with tetraploid (2n 5 48) and octoploid (2n 5 96)being the most frequently observed (Clevering and Lissner1999). Shoots of octoploid generally are longer and thickerand have larger leaves than those of tetraploid (Clevering et al.2001; Hanganu et al. 1999; Pauca-Comanescu et al. 1999).This relationship between euploidy level and morphology iscommon because the most immediate and universal effect ofpolyploidy is an increase in cell size. However, polyploidydoes not always lead to an overall increase in the plant size,

because a common effect of polyploidy also is a reduction inthe number of cell divisions during development (Stebbins1971). It has been suggested that genetic variation amongpopulations from different geographic regions has arisen as aresult of growth in different climatic conditions (Cleveringet al. 2001). Along a latitudinal gradient, gradual changeoccurs in day length, and the same happens in the amount ofsolar radiation and air temperature. Common reed popula-tions, originating from different geographic regions along alatitudinal gradient from Northern Sweden to Spain, differedin time of cessation of growth, shoot morphology, andbiomass allocation. Under the same environmental condi-tions, common reed originating from higher latitudes startedgrowing earlier than southern populations, but finished thegrowth early in the season. The southern populations failed tocomplete the whole growth cycle before the first frost and didnot develop mature seeds (Clevering et al. 2001).

Morphological and physiological traits often are influencedby the environment, but genetic markers are not affected bythe environmental factors, so they might present the mostreliable method of distinguishing different reed populations(Saltonstall 2003a). Different molecular techniques have beenemployed to detect clonal diversity in common reed, such asallozymes (Clevering et al. 2001; Clevering and Lissner 1999;Hauber et al. 1991; Pellegrin and Hauber 1999), randomamplified polymorphic DNAs (RAPDs) (Curn et al. 2007;Keller 2000; Koppitz and Kuhl 2000; Kuhl et al. 1999;Neuhaus et al. 1993), restriction fragment length polymor-phisms (RFLPs) (Koppitz et al. 1997; Saltonstall 2003b) andamplified fragment length polymorphisms (AFLPs) (Lamber-tini et al. 2006), microsatellite (SSR) analysis, and chloroplastDNA sequencing (Saltonstall 2002, 2003a). The random-amplified polymorphic DNA technique was developed in1990 (Williams et al. 1990): it has become recognized as amore accessible technique, and it is relatively low-cost andrequires very small quantities of genomic DNA (Ragot andHoisington 1993; Russell et al. 1997). It is a fairly simplemethod, and has been used in many studies for theclarification of nomenclature (Keil and Griffin 1994),identification of herbarium accessions (Khadari et al. 1995),or elucidation of genetic relationships (Russell et al. 1997).

The probability of success for management strategies willincrease with understanding of the nature and extent ofgenetic diversity among reed clones. If genetically diverse

DOI: 10.1614/WS-D-10-00163.1* Assistant Professor, Associate Professor, Associate Professor, Associate

Professor, and Professor, Department of Agronomy and Plant Breeding,University of Tehran, Karaj, Iran. Current address of first author: Islamic AzadUniversity, Science and Research Branch, Tehran, Iran. Corresponding author’sE-mail: [email protected] and [email protected].

Weed Science 2011 59:366–375

366 N Weed Science 59, July–September 2011

Iranian clones of common reed evolve, they might responddifferentially to weed management strategies, includingherbicides. Published studies of population genetic variationin common reed have focused on geographically localizedpopulations in Europe and the United States (Djebrouni1992; Hansen et al. 2007; Koppitz et al. 1997; Kuhl andNeuhaus 1993; McKee and Richards 1996; Neuhaus et al.1993; Pellegrin and Hauber 1999; Zeidler et al. 1994). Thisstudy investigates the genetic diversity and morphologicaldifferences among clones of common reed in Iran.

Materials and Methods

Plant Material. The rhizomes of 39 clones of common reedused in this study were collected from different regions of Iranin March 2005. The sampling areas ranged from 39u339N.,47u469E (Moghan) to 31u549N, 47u269E (Shoosh) (Figure 1).For the purpose of this study, these plant clones are identifiedby name and abbreviation of their location of origin (Table 1).The minimum distance between locations was more than10 km. At each location, more than 50 rhizome pieces werecollected, and the stand of common reed sampled wasconsidered to be one clone (rhizomes collected from a singleplant). A day after collection, the rhizomes were divided intosmall portions with two or three visible, well-formed buds andtransplanted into an experimental plot. Before transplanting,the soil was plowed and harrowed. After transplanting wascompleted, the plots were immediately irrigated.

The rhizomes of clones were grown in plots (3 m by 3 m,with 2-m spacing from the adjacent plot) at Research Farm ofPardise of Agriculture and Natural Science, University ofTehran, Karaj, Iran (35u449N, 51u109E, altitude 1300 m)using a randomized complete block design with threereplications. The soil was a fine sandy-loam. Culturaloperations, including frequent irrigation (every 2 d) andmanual elimination of other weeds were done during season.

Plots were watched closely and repeated efforts were made tomaintain the purity of individual clones. A 2-yr field studywas conducted using rhizomes and methods were the same inboth years.

In the first year, ambient climatic conditions were measuredat a nearby weather station; the average air temperatures inApril, May, June, July, August, September, and October were11, 17, 25, 25, 28, 25, and 19 C, respectively.

Morphological Study. At maximum biomass, the timewhen shoot length did not increase further, the ten tallestshoots per replicate were harvested by clipping them atground level. The shoots were wrapped in large plastic bagsbefore being transported to the laboratory (Hansen et al.2007). In order to facilitate comparison between the clones,special attention was paid to the tallest shoots within eachsample, which had originated from the apical rhizome buds.Therefore, the shoots were of similar age and, because theyall belonged to the first spring cohort of shoots, theyprobably were little-affected by intraspecific competition(Cosentino et al. 2006).

In the laboratory, the length of each shoot was measuredfrom the clipped base to the uppermost leaf. Basal diameter(cm) was measured between the two lowest nodes. Numberof internodes (per shoot) was counted before the shoot wasdivided into leaf, stem (with leaf sheath), and panicle. Paniclelength (cm) was determined for each shoot. Length andwidth (cm) of the sixth leaf above the ground was measuredfor each shoot. Leaf area (cm2) was measured with leaf areameter.1 Aboveground dry weight (g) was determined afterdrying to constant weight at 80 C for 48 h. Aboveground dryweight was obtained by adding the dry weights of the stem,leaf, and panicle. At the end of the growing season, thepercentage of flowering shoots was determined. Time ofpanicle appearance and senescence (days after planting) alsowere recorded.

Figure 1. The geographic origin of the 39 clones of common reed used in this study.

Diyanat et al.: Genetic diversity of common reed N 367

RAPD Assay. Common reed leaf samples were taken andstored at 220 C until preparation. DNA for polymerase chainreaction (PCR) assay was isolated according to the CTABmethod of Rogers and Bendich (1985) and stored in 13TRIS-EDTA buffer solution at 4 C. The RAPD analysis wasperformed using a set of 16 random primers2 (Table 2) on allcommon reed clones. PCR was carried out in a 25 ml reactionmixture containing 1.9 mM MgCl2, 0.5ml primer, 1.253PCR Buffer, 0.2 mM of each deoxyribonucleotide triphos-phate (dNTP), approximately 100 ng genomic DNA, and 1 UTaq DNA polymerase.3 Amplifications were performed in athermal cycler programmed as follows: 1 min at 92 C, 1 minat 35 C, 2 min at 72 C, and finally at 72 C for 4 min.Amplified products were separated by gel electrophoresis in1.5% agarose and tris-acetate-EDTA (TAE) buffer. Amplifi-cation products were separated in 1.5% agarose gels run in 13TAE buffer and detected by staining with ethidium bromide.RAPD fingerprints were amplified repeatedly (the same resultswere obtained in three independent PCR experiments). Theclear and distinct banding pattern indicated suitability of thismethod.

Statistical Analyses. Quantitative analyses of morphologicaltraits were carried out using SAS software.4 Pearson’scoefficients were used to determine the degree of associationsamong the traits. Euclidean distances were estimated for all

pairs of clones using standardized values of the traits and theformula

dxy~ffipXn

i~1~1(Xi{yi)

2, ½1�

where xi and yi are the ith characters measured on two clones andn is the number of characters (Romesburg 1984). The datamatrix usually is standardized to recast the units of measurementas dimensionless units by standardizing the data; all traitsbecome equally important in determining the distances (Manly1986). Euclidean distance coefficients measure the literaldistance between two objects when they are viewed as pointsin the two-dimensional space formed by their attributes(Romesburg 1984), in this case morphological traits. Clusteranalysis was then performed using the distance coefficients(Swofford and Olsen 1990). The contribution of the measuredmorphological traits to the overall differences between the cloneswas explored using principal components analysis (PCA).

For RAPD analysis, polymorphic RAPD fragments werescored as 1 for the presence and 0 for the absence of a DNAband for each clones. The data matrix was entered into theNTSYS program (Rohlf 1998) and analyzed using thequalitative routine to generate Dice similarity index as inNei and Li (1979). A dendrogram showing geneticrelationships of the 39 clones was constructed using theunweighted pair-group method with arithmetic averages(UPGMA).

Table 1. Collection codes and origin sites of sampled clones of common reed.

No. Origin site Province Codes Latitude N Longitude E

1 Moghan Ardebil A-MO1 39u339 47u4692 Moghan Ardebil A-MO2 39u309 47u449

3 Moghan Ardebil A-MO3 39u319 48u0194 Moghan Ardebil A-MO4 39u339 48u0395 Moghan Ardebil A-MO5 38u159 47u5496 Moghan Ardebil A-MO6 38u509 47u0497 Mesgaran Khorasan KO-ME 36u479 55u4398 Mashhad Khorasan KO-MA 36u209 55u069

9 Nazarie Khorasan KO-NA 36u089 55u25910 Dezfool Khozestan KZ-D1 32u159 48u26911 Dezfool Khozestan KZ-D2 32u149 48u27912 Ojirob Khozestan KZ-OJ 32u129 48u29913 Motahhari-shahrak Khozestan KZ-MO 32u169 48u22914 Shoosh Khozestan KZ-S1 31u549 47u26915 Shoosh Khozestan KZ-S2 31u459 46u359

16 Sarabeyavari Kermanshah KE-S1 34u299 46u56917 Sarabeyavari Kermanshah KE-S2 34u399 46u19918 Bisotoon Kermanshah KE-BI 34u229 47u26919 Mahidasht Kermanshah KE-MA 34u199 47u52920 Shahre-Ray Tehran T-SHR 35u339 51u22921 Shahrake-Sinamayi Tehran T-SHS 35u279 51u139

22 Varamin-Goltape Tehran T-VGO 35u039 51u30923 Varamin-Gharchek Tehran T-VG1 35u099 51u36924 Varamin-Gharchek Tehran T-VG2 35u139 51u40925 Varamin-Gharchek Tehran T-VG3 35u209 51u47926 Dolat Abad Tehran T-DO1 35u469 51u10927 Dolat Abad Tehran T-DO2 35u409 51u02928 Mohammad Shahr Tehran T-MO1 35u509 51u069

29 Mohammad Shahr Tehran T-MO2 35u579 51u14930 Beheshtmasoome Qom Q-BE1 34u309 51u16931 Beheshtmasoome Qom Q-BE2 34u399 51u18932 Beheshtmasoome Qom Q-BE3 34u599 51u30933 Sari Mazandaran MA-S1 36u379 52u56934 Sari Mazandaran MA-S2 36u299 53u049

35 Gorgan Golestan G-GR1 36u079 54u16936 Gonbad Golestan G-GO1 37u109 54u43937 Gonbad Golestan G-GO2 37u479 54u51938 Gorgan Golestan G-GR2 36u309 54u05939 Gorgan Golestan G-GR3 36u519 54u289


Results

Variability of Morphological Traits. The results showedthat clones of common reed were significantly different in allevaluated morphological traits (Table 3), suggesting thatselection for these traits could be possible.

When grown under the same environmental conditions,common reed originating from higher latitudes began theirgrowth earlier than those of southern clones, but finishedgrowth early in the season. Shoot length ranged from 157.0 to237.6 cm. The southern clones KZ-OJ and KZ-MO had the

Table 2. RAPD primers used, their sequence of nucleotides, and the percent of polymorphic bands produced by each primer.

Marker no. Primer name Nucleotide sequence 59R39 Number of total bands Percent of polymorphic bands

1 UBC 1 CCT GGG CTT C 15 922 UBC 3 CCT GGG CTT A 8 87.53 UBC5 CCT GGG TTC C 9 804 UBC9 CCT GCG CTT A 9 805 UBC 13 CCT GGG TGG A 9 73.36 UBC16 GGT GGC GGG A 8 887 UBC64 GAG GGC GGG A 9 858 UBC 66 GAG GGC GTG A 10 75.69 UBC76 GAG CAC CAG T 8 90

10 UBC77 GAG CAC CAG G 9 8011 UBC82 GGG CCC GAG G 8 7512 UBC 84 GGG CGC GAG T 10 9013 UBC89 GGG GGC TTG G 8 83.314 UBC95 GGG GGG TTG G 10 80.915 UBC96 GGC GGC ATG G 10 86.916 UBC100 ATC GGG TCC G 9 83.3Total 149Average 9.31 80.98

Table 3. Variation in different morphological traits among common reed clones (6 standard error).

Clone Shoot length (cm) Shoot diameter (cm)Number of internodes

(per shoot) Leaf length (cm) Leaf width (cm)

A-MO1 179.0 6 6.67 0.45 6 0.05 16.67 63.15 36.98 6 2.23 2.83 6 0.14A-MO2 185.3 6 6.33 0.48 6 0.04 16.33 62.37 36.42 6 2.25 3.03 6 0.17A-MO3 167.0 6 6.83 0.50 6 0.03 16.00 62.32 37.38 6 5.00 2.97 6 0.25A-MO4 170.3 6 7.77 0.47 6 0.02 16.67 62.32 39.80 6 5.25 2.87 6 0.21A-MO5 170.6 6 5.01 0.45 6 0.03 18.33 63.15 40.83 6 5.69 2.87 6 0.12A-MO6 172.3 6 8.10 0.51 6 0.04 16.67 63.89 39.83 6 4.55 2.80 6 0.42G-GO1 188.6 6 9.16 0.38 6 0.04 16.83 63.33 52.75 6 3.52 2.54 6 0.46G-GO2 192.0 6 8.81 0.41 6 0.04 18.50 61.33 49.73 6 7.88 2.70 6 0.18G-GR1 180.3 6 5.47 0.41 6 0.04 16.83 62.11 55.45 6 4.33 2.73 6 0.05G-GR2 178.0 6 8.26 0.38 6 0.04 18.33 63.44 52.42 6 2.52 2.30 6 0.19G-GR3 177.0 6 5.53 0.41 6 0.03 18.50 61.15 51.80 6 4.70 2.40 6 0.20KE-BI 176.6 6 8.24 0.45 6 0.02 15.00 61.50 40.45 6 3.61 2.54 6 0.45KE-MA 170.6 6 8.16 0.54 6 0.02 16.17 62.89 40.78 6 3.48 2.70 6 0.16KE-S1 181.0 6 8.54 0.50 6 0.04 14.00 62.37 42.10 6 4.33 3.21 6 0.29KE-S2 189.0 6 9.05 0.48 6 0.03 14.17 62.29 42.05 6 1.46 3.10 6 0.20KO-MA 157.0 6 4.50 0.32 6 0.02 15.00 62.80 40.08 6 2.07 2.21 6 0.39KO-NA 173.3 6 8.32 0.45 6 0.02 18.33 63.35 43.05 6 2.13 2.90 6 0.25KO-ME 161.0 6 3.41 0.30 6 0.04 14.17 62.03 38.82 6 4.06 2.14 6 0.11KZ-D1 206.0 6 8.68 0.45 6 0.03 21.00 62.67 52.45 6 3.74 3.21 6 0.50KZ-D2 214.6 6 4.33 0.45 6 0.02 21.17 62.89 50.68 6 2.64 3.34 6 0.44KZ-MO 230.3 6 9.47 0.53 6 0.05 21.83 62.48 51.62 6 5.08 3.60 6 0.18KZ-OJ 237.6 6 4.08 0.53 6 0.03 20.00 62.23 51.50 6 2.51 3.35 6 0.10KZ-S1 221.6 6 5.90 0.60 6 0.02 23.50 63.23 50.42 6 3.46 3.70 6 0.23KZ-S2 228.3 6 6.08 0.60 6 0.05 23.67 61.91 46.95 6 3.27 3.81 6 0.51MA-S1 171.0 6 6.51 0.60 6 0.03 20.17 63.58 36.75 6 3.05 4.21 6 0.27MA-S2 193.0 6 8.14 0.51 6 0.05 16.17 62.77 43.12 6 3.48 3.40 6 0.40Q-BE1 187.0 6 7.40 0.47 6 0.02 16.67 62.94 34.05 6 1.69 2.10 6 0.16Q-BE2 173.0 6 8.34 0.48 6 0.05 19.17 63.70 35.07 6 2.25 1.83 6 0.12Q-BE3 184.0 6 9.85 0.54 6 0.03 16.50 62.02 34.05 6 3.50 2.04 6 0.35T-DO1 185.6 6 5.40 0.45 6 0.05 14.17 62.47 43.40 6 4.15 3.64 6 0.45T-DO2 180.3 6 11.46 0.45 6 0.04 13.50 62.08 43.43 6 2.92 3.51 6 0.55T-MO1 195.0 6 9.84 0.48 6 0.04 15.00 62.82 44.75 6 2.88 3.13 6 0.10T-MO2 181.0 6 12.0 0.48 6 0.03 16.00 61.37 44.47 6 2.47 3.10 6 0.21T-SHR 195.0 6 9.93 0.42 6 0.03 15.17 61.54 44.82 6 4.24 3.11 6 0.32T-SHS 200.0 6 7.38 0.44 6 0.05 15.33 62.70 45.00 6 2.54 2.90 6 0.20T-VG1 190.6 6 6.86 0.50 6 0.02 15.83 61.91 43.78 6 2.71 3.23 6 0.25T-VG2 179.6 6 11.29 0.47 6 0.05 16.00 62.65 44.40 6 3.44 3.33 6 0.19T-VG3 184.3 6 4.82 0.44 6 0.03 15.17 63.40 44.98 6 2.94 3.30 6 0.29T-VGO 173.0 6 6.8 0.48 6 0.03 14.33 61.73 42.73 6 2.81 3.30 6 0.23LSD 9.32 0.02 3.12 4.53 0.33


longest shoots (237.6 and 230.3, respectively). The northernclone KO-MA had the shortest shoot (157.0 cm) followed byKO-ME with the value of 161.0 cm (Table 3). The basalshoot diameter varied from 0.32 (KO-ME) to 0.60 cm (KZ-S1, KZ-S2, and MA-S1). The highest and lowest number ofshoot internodes were observed in KZ-S2 (23.67) and T-DO2(13.5), respectively. The leaf length and leaf width rangedfrom 34.05 to 55.45 cm and from 1.83 to 4.21 cm,respectively. Moreover, the highest and lowest leaf lengthswere observed in G-GR1 and Q-BE1,and Q-BE3 respectively.MA-S1 had the highest and Q-BE2 had the lowest leaf widths(Table 3). The leaf area per shoot ranged from 664 cm2 (KZ-D2) to 321 cm2 (KO-ME and KE-S1) (Table 4). The highestaboveground dry weight per shoot was observed in KZ-OJ(12.83 g) whereas KE-S1 had the lowest aboveground dryweight per shoot (7.86 g). The results obtained showed thatthe plants of clones from the southern areas were taller andthicker with more nodes, and they also were more vigorousthan plants from more northern areas. The percentage offlowering shoots at the end of growing season was rangedfrom 15.0 (KE-S2) to 95.0 (MA-S1). In all clones, the time ofpanicle appearance was negatively correlated with latitude oforigin (Figure 2). The range of panicle length was from23.00 cm (KZ-OJ) to 39.17 cm (MA-S1). The panicles were

larger in southern clones than those of northern clones. Thesenescence time varied among clones, with the most strikingdifference being A-MO3 that started senescing in July, andKZ-S2 which started senescing in October (Table 4).

The phenotypic correlation between traits was shown inTable 5. Shoot length was significantly and positivelycorrelated with all traits except percentage of floweringshoots. The strongest relationship was between shoot lengthand aboveground dry weight (r2 5 0.88). Leaf length also was

Figure 2. Relationship between the latitude of origin and time of panicleappearance for clones of common reed.

Table 4. Variation in different morphological traits among common reed clones (6 standard error).

CloneLeaf area (per plant)

(cm2)Aboveground dry

weight (g)Percentage of flowering

shoots Panicle length (cm)Number of days until

panicle appearanceNumber of days untilsenescence appearance

A-MO1 571.0 6 11.83 9.62 6 0.55 85.0 6 1.61 30.17 6 3.71 144.77 6 6.23 160.00 6 3.14A-MO2 556.0 6 15.22 9.66 6 1.43 84.0 6 1.80 31.17 6 4.08 140.38 6 3.99 156.00 6 3.29A-MO3 563. 0 6 16.04 9.48 6 0.68 89.0 6 1.89 34.17 6 4.62 139.22 6 5.08 155.00 6 3.25A-MO4 539.0 6 6.45 9.51 6 0.92 90.0 6 0.76 35.00 6 5.77 142.05 6 4.97 158.00 6 3.80A-MO5 558.0 6 15.22 9.93 6 0.93 80.0 6 2.08 35.83 6 5.35 146.48 6 3.79 162.00 6 4.32A-MO6 560.0 6 28.35 9.74 6 1.56 82.0 6 0.76 35.83 6 3.33 143.58 6 4.08 159.00 6 4.00G-GO1 522.0 6 15.22 9.27 6 0.45 70.0 6 5.51 31.33 6 5.55 162.57 6 5.60 179.17 6 4.36G-GO2 529.0 6 24.64 10.35 6 0.61 74.0 6 3.28 31.17 6 6.57 161.37 6 5.02 178.00 6 6.87G-GR1 520.0 6 9.31 8.48 6 0.60 69.0 6 3.91 30.00 6 3.80 169.33 6 3.85 186.17 6 3.99G-GR2 538.0 6 12.99 9.02 6 1.26 75.0 6 1.04 32.17 6 3.08 165.28 6 5.09 177.33 6 6.37G-GR3 531.0 6 24.39 9.07 6 1.64 80.0 6 1.53 31.17 6 4.06 162.13 6 4.39 180.17 6 2.47KE-BI 329.0 6 16.64 8.27 6 0.65 17.0 6 1.04 26.17 6 5.28 165.22 6 3.46 182.17 6 2.60KE-MA 334.0 6 27.17 8.09 6 0.49 29.0 6 0.87 27.00 6 6.32 163.52 6 3.89 180.17 6 7.75KE-S1 321.0 6 6.23 7.86 6 0.87 20.0 6 1.04 24.17 6 5.33 170.37 6 4.32 185.50 6 2.64KE-S2 333.0 6 15.76 8.01 6 0.72 15.0 6 0.87 25.00 6 2.23 172.18 6 1.83 189.17 6 2.93KO-MA 330.0 6 22.97 8.57 6 0.61 75.0 6 7.55 34.33 6 6.25 150.32 6 2.63 172.17 6 7.46KO-NA 361.0 6 28.30 8.93 6 0.25 76.0 6 1.61 33.33 6 4.73 153.50 6 2.88 176.67 6 4.01KO-ME 321.0 6 5.12 8.62 6 0.38 85.0 6 1.76 34.00 6 4.18 155.27 6 7.52 166.60 6 5.42KZ-D1 641.0 6 10.49 11.56 6 0.26 30.0 6 3.46 30.17 6 6.38 190.35 6 5.80 208.17 6 3.34KZ-D2 664.0 6 7.78 12.23 6 0.42 42.0 6 2.78 24.00 6 3.05 192.37 6 3.59 209.83 6 5.76KZ-MO 619.0 6 15.64 12.79 6 0.50 24.0 6 2.47 24.00 6 2.32 197.58 6 2.65 215.17 6 5.22KZ-OJ 621.0 6 15.45 12.83 6 0.53 33.67 6 2.02 23.00 6 2.00 196.53 6 5.61 214.00 6 3.62KZ-S1 595.0 6 6.27 12.59 6 1.43 20.33 6 3.21 26.33 6 7.25 200.28 6 2.81 218.33 6 6.11KZ-S2 585.0 6 11.56 12.57 6 0.57 19.0 6 3.12 27.17 6 2.12 203.93 6 4.54 222.50 6 3.51MA-S1 486.0 6 9.69 9.38 6 0.95 95.0 6 3.50 39.17 6 6.36 159.38 6 3.07 176.17 6 5.62MA-S2 521.0 6 13.32 9.42 6 1.22 80.0 6 1.32 33.17 6 2.75 158.23 6 7.12 175.00 6 5.50Q-BE1 443.0 6 13.49 8.36 6 0.83 90.0 6 2.50 28.00 6 3.13 172.32 6 3.34 189.00 6 6.25Q-BE2 641.0 6 11.76 8.48 6 0.35 80.0 6 5.89 30.17 6 3.33 177.58 6 4.19 193.00 6 4.77Q-BE3 463.0 6 6.07 8.83 6 1.09 85.0 6 4.92 31.17 6 4.25 179.40 6 3.20 197.00 6 4.17T-DO1 440.0 6 9.94 8.36 6 0.78 70.0 6 3.50 30.33 6 3.84 168.57 6 5.09 185.00 6 8.67T-DO2 431.0 6 28.66 8.53 6 0.61 65.0 6 0.29 32.33 6 2.85 169.37 6 3.24 186.00 6 3.92T-MO1 465.0 6 16.93 8.43 6 0.62 60.0 6 5.06 29.17 6 2.31 168.33 6 3.42 185.17 6 4.52T-MO2 476.0 6 4.36 8.46 6 1.08 62.0 6 3.50 30.00 6 4.58 167.37 6 7.28 184.17 6 4.42T-SHR 497.0 6 10.90 9.53 6 1.02 80.0 6 5.29 25.33 6 5.12 170.83 6 4.83 187.00 6 5.33T-SHS 484.0 6 16.88 9.38 6 .066 82.0 6 4.33 28.17 6 4.69 171.60 6 5.63 188.17 6 7.91T-VG1 484.0 6 18.95 8.74 6 .036 75.0 6 0.50 27.00 6 3.91 173.33 6 2.78 190.00 6 4.41T-VG2 467.0 6 9.22 8.48 6 0.65 70.0 6 4.75 31.00 6 3.31 172.75 6 4.35 189.17 6 3.40T-VG3 323.0 6 9.18 8.38 6 0.43 71.0 6 3.91 29.00 6 2.30 170.25 6 4.36 187.17 6 5.79T-VGO 497.0 6 4.44 8.57 6 1.31 80.0 6 4.09 33.00 6 4.18 173.45 6 5.18 190.00 6 4.47LSD 18.45 1.01 7.29 5.27 5.46 5.97


positively correlated with shoot diameter, leaf area, above-ground dry weight, panicle length, time of panicle appearance,and senescence time, but negatively correlated with thepercentage of flowering shoots and number of internodes. Thepercentage of flowering shoots and leaf area also exhibitednegative correlation with each other.

Clustering of common reed clones on the basis of distancevalues for morphological traits produced a dendrogram withfour major groups (Figure 3). The first group was comprisedof six clones form Ardebil (A-MO1, A-MO2, A-MO3,A-MO4, A-MO5, and A-MO6), one population fromMazandaran (MA-S1), and five clones from Golestan(G-GR1, G-GR2, G-GR3, G-GO1, and G-GO2). The

Table 5. Simple correlation coefficients among morphological traits in 39 clones of common reed.

Shootlength

Leaflength

Leafwidth

Shootdiameter

Number ofinternodes

Leafarea

Abovegrounddry weight

Percentage offlowering

shootsPaniclelength

Number ofdays until

panicleappearance

Number ofdays untilsenescenceappearance

Shoot length 1.00Leaf length 0.45** 1.00Leaf width 0.40* 0.25 1.00Shoot diameter 0.69** 0.41** 0.24 1.00Number of

internodes 0.45** 20.16 0.57** 20.06 1.00Leaf area 0.67** 0.39* 0.30 0.01 0.64** 1.00Aboveground dry

weight 0.88** 0.44** 0.41** 20.22 0.82** 0.42** 1.00Percentage of

flowering shoots 20.84** 20.41** 0.42** 0.22 0.79** 20.39* 0.69** 1.00Panicle length 0.32* 0.35* 0.37* 0.23 0.38* 0.32* 0.31* 0.24 1.00Number of days until

panicle appearance 0.41** 0.39* 0.47** 0.23 0.47** 0.36* 0.35* 0.34* 0.37* 1.00Number of days until

senescence appearance 0.65** 0.46** 0.34* 0.01 0.54** 0.40* 0.22 0.57** 0.60** 0.69** 1.00

**’* Significant at 1% and 5%, respectively.

r

Figure 3. Dendrogram generated for clones of common reed on the basis ofunweighted pair group method with arithmetic averages (UPGMA) usingmorphological traits data from 2 yr. Scale represents Euclidean distance value (d ).

Figure 4. Principal components analysis (PCA) scores of clones of common reedcalculated from morphological data.


second group included ten clones from Tehran (T-SHR, T-SHS, T-VGO, T-VG1, T-VG2, T-VG3, T-DO1, T-DO2,T-MO1, and T-MO2), three clones from Qom (Q-BE1,Q-BE2, and Q-BE3), and one clone from Mazandaran(MA-S2). The third group contained six clones fromKhozestan (KZ-SH, KZ-OJ, KZ-D1, KZ-D2, KZ-A1, andKZ-A2). The fourth group comprised three clones fromKhorasan (KO-MA, KO-ME, and KO-NA) and four clonesfrom Kermanshah (KE-S1, KE-S2, KE-BI, and KE-MA).What we can conclude from this dendrogram is the clearseparation of the clones based on location (province), with theexception of the clones from Mazandaran.

In the PCA analysis, the morphological traits of clones didnot discriminate well between clones (Figure 4) in agreementwith Curn et al (2007). The first and second PCA axes ofmorphological traits accounted for 49.29% and 18.57% oftotal variation, respectively (Figure 4). The traits containingaboveground dry weight, number of internodes, leaf area, andshoot length explained variability associated with the first axis.The panicle length, number of days until panicle appearance,and senescence explained variability associated with thesecond axis.

RAPD Analysis. A total of 149 bands were screened, amongwhich 123 were polymorphic. The number of bands per

primer varied from 8 to 15 with an average of 9.31. Theaverage proportion of polymorphic markers across primerswas 83.1%, ranging between 73.3% (UBC13) and 92%(UBC1) (Table 2).

Estimates of genetic similarity of RAPD based on 123.1polymorphic markers among 39 clones of common reedranged from 0.43 to 0.85 with an average of 0.60 (Table 6).Maximum similarity was with T-MO1 and T-MO2 (0.85);minimum similarity was with MA-S1 and G-GR1 (0.43).

Cluster analysis resulted in grouping of the 39 clones ofcommon reed into four main clusters in 0.62 distance unit(Figure 5). The first cluster had two subdivisions. The firstsubdivision contained six clones form Ardebil (A-MO1,A-MO2, A-MO3, A-MO4, A-MO5, and A-MO6), threeclones from Khorasan (KO-MA, KO-ME, and KO-NA), andfour clones form Kermanshah (KE-S1, KE-S2, KE-BI, andKE-MA). The second subdivision contained six clones fromKhozestan (KZ-SH, KZ-OJ, KZ-D1, KZ-D2, KZ-A1, andKZ-A2), one clone from Mazandaran (MA-S2), and 10 clonesfrom Tehran (T-SHR, T-SHS, T-VGO, T-VG1, T-VG2,T-VG3, T-DO1, T-DO2, T-MO1, and T-MO2). Thesecond cluster included clones from Qom (Q-BE1, Q-BE2,and Q-BE3). Two clones from Golestan (G-GO1 andG-GO2) form the third cluster, and three clones from theGolestan (G-GR1, G-GR2, and G-GR3) formed the fourth

Table 6. Similarity matrix among common reed clones using Nei and Li’s coefficient based on RAPD bands.

A-MO1 A-MO2 A-MO3 A-MO4 A-MO5 A-MO6 KZ-D1 KZ-D2 KZ-OJKZ-MO

KZ-S1 KZ-S2 MA-S1 MA-S2 KE-S1 KE-S2 KE-BI KE-MA

A-MO1 1.00A-MO2 0.78 1.00A-MO3 0.76 0.81 1.00A-MO4 0.72 0.69 0.67 1.00A-MO5 0.73 0.69 0.64 0.74 1.00A-MO6 0.73 0.74 0.67 0.76 0.75 1.00KZ-D1 0.61 0.56 0.61 0.68 0.60 0.56 1.00KZ-D2 0.62 0.55 0.61 0.68 0.61 0.60 0.84 1.00KZ-OJ 0.62 0.61 0.55 0.67 0.65 0.62 0.70 0.72 1.00KZ-MO 0.70 0.65 0.58 0.74 0.71 0.68 0.72 0.71 0.83 1.00KZ-S1 0.62 0.61 0.61 0.56 0.55 0.55 0.72 0.71 0.69 0.66 1.00KZ-S2 0.65 0.64 0.59 0.66 0.64 0.60 0.69 0.72 0.70 0.75 0.71 1.00MA-S1 0.49 0.48 0.52 0.58 0.50 0.47 0.66 0.57 0.59 0.55 0.59 0.62 1.00MA-S2 0.57 0.55 0.51 0.63 0.52 0.61 0.59 0.62 0.60 0.64 0.58 0.63 0.45 1.00KE-S1 0.64 0.69 0.57 0.62 0.61 0.62 0.56 0.53 0.68 0.72 0.66 0.62 0.55 0.62 1.00KE-S2 0.69 0.68 0.61 0.67 0.65 0.69 0.62 0.62 0.69 0.68 0.59 0.63 0.57 0.55 0.66 1.00KE-BI 0.59 0.64 0.57 0.61 0.58 0.53 0.63 0.64 0.62 0.64 0.71 0.68 0.54 0.61 0.58 0.56 1.00KE-MA 0.66 0.70 0.58 0.69 0.63 0.62 0.62 0.57 0.63 0.67 0.66 0.67 0.53 0.60 0.65 0.68 0.83 1.00Q-BE1 0.64 0.60 0.56 0.53 0.63 0.59 0.55 0.58 0.60 0.63 0.58 0.58 0.49 0.48 0.65 0.66 0.53 0.51Q-BE2 0.67 0.64 0.58 0.63 0.58 0.61 0.59 0.63 0.65 0.65 0.63 0.63 0.56 0.54 0.65 0.65 0.59 0.60Q-BE3 0.67 0.62 0.63 0.55 0.64 0.59 0.65 0.61 0.65 0.66 0.63 0.65 0.57 0.51 0.64 0.65 0.63 0.60T-SHR 0.62 0.57 0.60 0.64 0.59 0.57 0.69 0.71 0.68 0.67 0.71 0.72 0.63 0.62 0.59 0.63 0.61 0.61T-SHS 0.59 0.54 0.54 0.66 0.64 0.57 0.68 0.70 0.67 0.69 0.61 0.66 0.54 0.64 0.62 0.59 0.55 0.58T-VGO 0.57 0.54 0.54 0.66 0.58 0.57 0.70 0.68 0.68 0.65 0.65 0.68 0.54 0.61 0.60 0.65 0.55 0.58T-VG1 0.61 0.62 0.56 0.59 0.56 0.59 0.59 0.59 0.61 0.64 0.70 0.71 0.58 0.61 0.56 0.61 0.67 0.65T-VG2 0.61 0.60 0.58 0.64 0.54 0.61 0.63 0.65 0.58 0.64 0.67 0.69 0.56 0.61 0.60 0.61 0.59 0.58T-VG3 0.54 0.53 0.48 0.61 0.57 0.56 0.58 0.53 0.55 0.59 0.66 0.67 0.53 0.63 0.58 0.65 0.67 0.68T-DO1 0.66 0.71 0.64 0.59 0.70 0.62 0.60 0.56 0.68 0.75 0.60 0.65 0.54 0.56 0.69 0.64 0.58 0.61T-DO2 0.63 0.60 0.61 0.67 0.63 0.65 0.67 0.64 0.73 0.72 0.65 0.63 0.60 0.65 0.62 0.60 0.60 0.64T-MO1 0.64 0.61 0.58 0.64 0.60 0.62 0.61 0.63 0.66 0.70 0.63 0.70 0.52 0.69 0.59 0.58 0.66 0.63T-MO2 0.66 0.63 0.58 0.66 0.60 0.64 0.66 0.64 0.69 0.75 0.69 0.70 0.54 0.69 0.61 0.64 0.68 0.68KO-ME 0.65 0.63 0.54 0.64 0.62 0.59 0.63 0.58 0.63 0.67 0.56 0.64 0.56 0.61 0.69 0.77 0.63 0.69KO-MA 0.64 0.59 0.52 0.64 0.59 0.60 0.62 0.56 0.61 0.68 0.63 0.69 0.57 0.60 0.67 0.68 0.70 0.70KO-NA 0.65 0.64 0.57 0.69 0.59 0.63 0.58 0.55 0.62 0.66 0.57 0.69 0.53 0.65 0.66 0.70 0.65 0.69G-GR1 0.64 0.61 0.61 0.65 0.57 0.60 0.67 0.70 0.68 0.70 0.66 0.65 0.43 0.58 0.57 0.58 0.58 0.59G-GO1 0.59 0.61 0.58 0.66 0.55 0.67 0.62 0.65 0.72 0.69 0.58 0.61 0.47 0.64 0.67 0.65 0.51 0.58G-GO2 0.54 0.55 0.53 0.61 0.55 0.61 0.51 0.50 0.71 0.66 0.56 0.54 0.51 0.53 0.68 0.63 0.57 0.58G-GR2 0.63 0.65 0.60 0.64 0.60 0.65 0.61 0.58 0.72 0.73 0.58 0.63 0.49 0.55 0.65 0.68 0.53 0.58G-GR3 0.56 0.61 0.55 0.58 0.59 0.62 0.55 0.55 0.72 0.70 0.63 0.62 0.51 0.53 0.67 0.64 0.56 0.63


cluster. The fifth cluster contained only one clone fromMazandaran (MA-S1).

Another interesting attempt was the comparison of geneticsimilarities within provinces. The geographic distance be-tween KZ-D1 and KZ-D2 was less than 12 km and geneticsimilarity between these clones was 0.84. All clones fromQom (Q-BE1, Q-BE2, and Q-BE3) were placed in the samecluster. Genetic distances among these clones were lessbecause Qom is a small province with a low variability inclimatic conditions. Similarly, genetic distances among clonesfrom Ardrbil (A-MO1, A-MO2, A-MO3, A-MO4, A-MO5,and A-MO6) were small. Genetic distances among clonesfrom Tehran were large; this might be because of variableclimatic conditions in Tehran. T-SHR and T-SHS cloneswere from the south but T-DO1, T-DO2, T-MO1, andT-MO2 clones were from northwest, and T-VGO, T-VG1,T-VG2, and T-VG3 clones were from southeastern ofTehran.

Discussion

The genetic diversity in clonal plant population has beenstudied by some researchers, and they found substantialamounts of diversity in most plant species (Diggle et al.1998; Ellstrand and Roose 1987; Khudamrongsawart et al.

Figure 5. Dendrogram of individual clones of common reed constructed on thebasis of RAPD data, by unweighted pair group method with arithmetic averages(UPGMA) method, resulting from similarity matrix calculated with the metric ofNei and Li (1979).

Q-BE1

Q-BE2

Q-BE3

T-SHR

T-SHS

T-VGO

T-VG1

T-VG2

T-VG3

T-DO1

T-DO2

T-MO1

T-MO2

KO-ME

KO-MA

KO-NA

G-GR1

G-GO1

G-GO2

G-GR2

G-GR3

1.000.76 1.000.80 0.71 1.000.57 0.60 0.68 1.000.53 0.55 0.63 0.80 1.000.59 0.62 0.54 0.64 0.60 1.000.55 0.62 0.60 0.64 0.64 0.64 1.000.55 0.63 0.53 0.65 0.66 0.64 0.79 1.000.56 0.59 0.59 0.61 0.61 0.63 0.76 0.70 1.000.64 0.67 0.70 0.57 0.61 0.65 0.65 0.59 0.56 1.000.57 0.63 0.63 0.66 0.65 0.74 0.65 0.67 0.62 0.78 1.000.59 0.67 0.62 0.58 0.58 0.65 0.64 0.64 0.61 0.72 0.73 1.000.63 0.69 0.64 0.65 0.62 0.69 0.71 0.67 0.66 0.70 0.73 0.85 1.000.57 0.61 0.60 0.62 0.58 0.57 0.62 0.62 0.69 0.59 0.57 0.55 0.62 1.000.61 0.62 0.66 0.63 0.62 0.53 0.65 0.64 0.68 0.59 0.62 0.60 0.68 0.84 1.000.52 0.64 0.57 0.59 0.51 0.59 0.64 0.60 0.62 0.60 0.56 0.64 0.64 0.76 0.70 1.000.51 0.58 0.62 0.61 0.55 0.55 0.53 0.50 0.47 0.58 0.58 0.59 0.59 0.55 0.59 0.62 1.000.57 0.58 0.58 0.64 0.56 0.56 0.52 0.51 0.51 0.57 0.57 0.55 0.60 0.60 0.58 0.66 0.71 1.000.52 0.55 0.60 0.56 0.56 0.58 0.54 0.53 0.55 0.59 0.59 0.58 0.62 0.58 0.60 0.65 0.57 0.62 1.000.57 0.62 0.58 0.64 0.58 0.68 0.57 0.59 0.56 0.65 0.65 0.62 0.64 0.58 0.56 0.63 0.59 0.68 0.73 1.000.58 0.64 0.64 0.52 0.52 0.60 0.62 0.58 0.60 0.67 0.64 0.67 0.63 0.58 0.57 0.59 0.55 0.59 0.70 0.75 1.00

Table 6. Extended.


2004; Widen et al. 1994). European studies of common reed,examining genetic variation and population genetic structure,have shown high levels of genetic divergence among popula-tions (Koppitz et al. 1997; Kuhl and Neuhaus 1993; Zeidler etal. 1994), both homogenous (Koppitz et al. 1997) orheterogenous populations (McKee and Richards 1996), or acombination of the two (Kuhl and Neuhaus 1993; Zeidler et al.1994). The high degree of genetic diversity detected amongreed populations in this study agrees with European studies.McLellan et al. (1997) stated that gene flow by means ofdispersal of vegetative propagules plays an important role inproviding variation among or within clonal populations.

Our results are in agreement with Bastlova et al. (2006),who showed that the clones from the southern areas grewtaller and more vigorously than plants from more northernareas. The clones from southern areas allocated the reduceddry mass proportion to generative reproduction. The resultsare in agreement with Clevering et al. (2001), who showedthat differences in length of growing season, time of flowering,morphology, and biomass allocation were sustained whenclones of common reed from different geographic origins weretransplanted to common environments. Karunaratne andAsaeda (2002) have shown that rhizome biomass and rhizomestanding stocks of nonstructural carbohydrates and mineralnutrients decrease in the early growing season and increaselater in the year in common reed. Storage is especiallyimportant in perennial plants living in regions with coldwinters, because the nonstructural carbohydrates are involvedin the tolerance of cold climatic conditions, including thedanger of frost damage and prolonged winters followed byvegetative periods with low temperatures, which limits carbonassimilation (Klimes et al. 1999). Therefore, the commonreed clones in northern regions are not as productive as thosein warmer lower latitudes. Weber and Schimd (1998) alsoshowed populations of two goldenrod (Solidago) speciesoriginating from northern locations flowered earlier andreached a smaller size at maturity than plants from southernlocations. Photoperiod, a factor of great importance, is highlyvariable across latitude gradient and influences plant life-history traits. Variability in phenology found in our studymight be strongly correlated with physiological requirementfor floral initiation and probably is genetically based.

RAPD is an effective method to detect intra- andinterpopulation variation and still is used widely for thesepurposes in many plants (Bussell et al. 2005; Curn et al. 2007;Keller 2000; Koppitz 1999; Koppitz et al. 1997; Kuhl et al.1999). Our results also show that RAPD is suitable for geneticdiversity assessment in reed clones.

In common reed research, the existence of ecotypes has beenmentioned frequently. However, in most instances, commonreed clones have not been grown under similar (controlled)conditions. Therefore, it is hard to tell whether differencesfound between populations or clones are due to environmen-tally induced variation rather than to genetic differences. In thisstudy we investigated genetic diversity in 39 Iranian clones ofcommon reed based on RAPD markers and morphologicaltraits. For this study, plants were grown under similarenvironmental conditions in terms of soil, water, nutrients,and climate. It is, therefore, fair to believe that any differencesin morphology traits are genetically determined. Clevering et al.(2001) also showed that differences in length of growing season,time of flowering, morphology, and biomass allocation were

sustained when genotypes of common reed from differentgeographic origins were transplanted to common environ-ments. Genetics and environments both play a role in theexpression of common reed characteristics. Cluster analysisbased on morphological traits grouped clones better thanRAPD markers, this possibly could be explained by the fact thatmorphological traits provided a better genome-wide coveragethan RAPD markers. Possibly, these variations among cloneswill assist in successful management of common reed usingchemical or other methods of control.

Sources of Materials

1 Leaf area meter, Li-Cor, Inc., Lincoln, NE.2 Sixteen random primers, University of British Colombia,

Vancouver, Canada.3 MgCl2 (50 Mm), 103 PCR Buffer, dNTPs Mix (10 mM), Taq

DNA polymerase (100U [5u/ml]), CinnaGen, Co., Tehran, Iran.4 Statistical software, Statistical Analysis Systems (version 9), SAS

Institute, Inc., 100 SAS Campus Drive, Cary, NC 27513.

Literature Cited

Bastlova, D., M. Bastl, H. Cızkova, and J. Kvet. 2006. Plasticity of Lythrumsalicaria and Phragmites australis growth chrarateristic across a Europeangeographical gradient. Hydrobiologia 570:237–242.

Bjork, S. 1967. Ecological investigation in Phragmites communis: studies intheoretic and applied limnology. Folia Limnol. Scand. 14:1–248.

Bussell, G. D., M. Waycott, and J. A. Chappill. 2005. Arbitrarily amplified DNAmarkers as characters for phytogenetic interference. Perspect. Plant Ecol. Evol.Syst. 7:3–26.

Clevering, O. A. 1999. Between- and within-population differences in Phragmitesaustralis. I. The effects of nutrient on seedling growth. Oecologia 121:447–457.

Clevering, O. A., H. Brix, and J. Lukaska. 2001. Geographic variation in growthresponse in Phragmites australis. Aquat. Bot. 69:89–108.

Clevering, O. A. and J. Lissner. 1999. Taxonomy, chromosome numbers, clonaldiversity and population dynamics of Phragmites australis. Aquat. Bot. 64:185–208.

Cosentino, S. L., V. Copani, G. M. Agosta, E. Sanzone, and M. Mantineo. 2006.First results on evaluation of Arundo donax L. clones collected in southernItaly. Ind. Crops Prod. 23:212–222.

Curn, V., B. Kubatova, P. Vavrova, O. Krivackova-Sucha, and H. Cizkova. 2007.Phenotypic and genotypic variation of Phragmites australis: comparison ofpopulations in two human-made lakes of different age and history. Aquat. Bot.86:321–330.

de Kroon, H. and J. Van Groenendael. 1997. The Ecology and Evolution ofGrowth in Clonal Plants. Leiden, The Netherlands: Backhuys Publishers. 453 p.

Diggle, P. K., S. Lower, and T. A. Runker. 1998. Clonal diversity in alpine populationsof Polygonum viviparum (Polygonaceae). Int. J. Plant Sci. 159:606–615.

Djebrouni, M. 1992. Variabilite, morphologique, caryologique et enzymatiguechez quelques populations de Phragmites austradis (Cav) Trin. ex Steud. FoliaGeobot. Phytotax. Praha. 27:49–59.

Dong, M. 1996. Clonal growth in plants in relation to resource heterogeneity:foraging behavior. Acta Bot. Sin. 30:828–835.

Ellstrand, N. C. and M. L. Roose. 1987. Patterns of genotypic diversity in clonalplant species. Am. J. Bot. 74:123–131.

Hanganu, J., M. Gridin, and H. Coops. 1999. Responses of ecotypes Phragmitesaustralis to increased seawater influence: a field study in the Danube Delta,Romania. Aquat. Bot. 64:351–358.

Hansen, D. L., C. Lambertini, A. Jampeetong, and H. Brix. 2007. Clone-specificdifferences in Phragmites australis: effects of ploidy level and geographic origin.Aquat. Bot. 86:269–279.

Hauber, D. P., D. A. White, S. P. Powers, and F. R. De Francesch. 1991.Isozyme variation and correspondence with unusual reflectance patterns inPhragmites australis (Poaceae). Plant Syst. Evol. 178:1–8.

Jackson, J.B.C., L. W. Buss, and R. E. Cook, eds. 1985. Population Biology andEvolution of Clonal Organisms. New Haven, CT, and London: YaleUniversity Press. 530 p.

Karunaratne, S. and T. Asaeda. 2002. Mathematical modeling as tool in aquaticecosystem management. J. Environ. Eng. 128:325–359.


Keil, M. and A. R. Griffin. 1994. Use of random amplified polymorphic DNA(RAPD) markers in the discrimination and verification of genotypes inEucalyptus. Theor. Appl. Genet. 88:442–450.

Keller, B.E.M. 2000. Genetic variation among and within populations ofPhragmites australis in the Charles River watershed. Aquat. Bot. 66:195–208.

Khadari, B., P. Lashermes, and F. Kjellberg. 1995. RAPD fingerprints foridentification and genetic characterization of fig (Ficus carica L.) genotypes.J. Genet. Breed. 49:77–86.

Khudamrongswart, J., R. Tayyar, and Y. S. Holt. 2004. Genetic diversity of giantreed (Arundo donax) in the Santa Ana River, California. Weed Sci. 52:395–405.

Klimes, L., J. Klimesova, and H. Cızkova. 1999. Carbohydrate storage inrhizomes of Phragmites australis (Cav.) Trin. ex Steud.: the effects of altitudeand rhizome age. Aquat. Bot. 64:105–110.

Koppitz, H. 1999. Analysis of genetic diversity among selected populations ofPhragmites australis world-wide. Aquat. Bot. 64:209–221.

Koppitz, H. and H. Kuhl. 2000. To the importance of genetic diversity of Phragmitesaustralis in the development of reed stands. Wetland Ecol. Manag. 8:403–414.

Koppitz, H., H. Kuhl, K. Hesse, and J.-G. Kohl. 1997. Some aspects of theimportance of genetic diversity in Phragmites australis (Cav.) Trin. ex Steudelfor the development of reed stands. Bot. Acta 110:217–223.

Kuhl, H., H. Koppitz, H. Rolletschek, and J.-G. Kohl. 1999. Clone specificdifferences in Phragmites australis stands. I. Morphology, genetic and sitedescription. Aquat. Bot. 64:235–246.

Kuhl, H. and D. Neuhaus. 1993. The genetic variability of Phragmites australisinvestigated by random amplified polymorphic DNA. Seeuferzerstorung andSeeferrenturierung in Mitteleeuropa-Limnologie aktuell. 5:9–18.

Lambertini, C., M.H.G. Gustafsson, J. Frydenberg, J. Lissneor, M. Speranza, andH. Brix. 2006. A polymorphic study of the cosmopolitan genus Phragmites(Poaceae) based on AFLP. Plant Syst. Evol. 258:161–182.

Manly, B.F.J. 1986. Multivariate Statistical Methods. London: Chapman andHall. 214 p.

McKee, J. and J. Richards. 1996. Variation in seed production and germinabilityin common reed (Phragmites australis) in Britain and France with respect toclimate. New Phytol. 133:233–24.

McLellan, A. J., D. Prati, O. Kaltz, and B. Schimd. 1997. Structure and analysisof phenotypic and genetic variation in clonal plants. Pages 185–210 in H. deKroon and J. van Groenendael, eds. The Ecology and Evolution of Growth inClonal Plants. Leiden, The Netherlands: Backhuys Publishers.

McNaughton, S. J. 1975. R- and K-Selection in Typha. Am. Nat. 109:251–261.Nei, M. and H. Li. 1979. Mathematical model for studying genetic variation in terms

of restriction endonucleuses. Proc. Natl. Acad. Sci. U. S. A. 79:5269–5273.Neuhaus, D., H. Kuhl, J.-G. Kohl, P. Dorfel, and T. Borner. 1993. Investigation

of genetic diversity of Phragmites stands using genomic fingerprinting. Aquat.Bot. 45:357–364.

Pauca-Comanescu, M., O. A. Clevering, J. Hanganu, and M. Gridin. 1999.Phenotypic differences among ploidy levels of Phragmites australis growing inRomania. Aquat. Bot. 64:223–234.

Pellegrin, D. and D. P. Hauber. 1999. Isozyme variation among populations of theclonal species, Phragmites australis (Cav). Trin. ex Steudel. Aquat. Bot. 63:241–259.

Ragot, M. and D. A. Hoisington. 1993. Molecular markers for plant breeding:comparison of RFLP and RAPD genotyping costs. Theor. Appl. Genet.86:975–984.

Raybould, A. F., A. J. Gray, M. J. Lawrence, and D. F. Marshall. 1991. Theevolution of Spartina G. E. Hubbard (Gramineae): origin and geneticvariability. Biol. J. Linn. Soc. 43:111–126.

Rogers, S. O. and A. J. Bendich. 1985. Extraction of DNA from milligram amountsof fresh, herbarium, and mummified plant tissues. Plant Mol. Biol. 5:69–76.

Rohlf, F. J. 1998. NTSYS–pc. Numerical Taxonomy and Multivariate AnalysisSystem Version 2.00. Setauket, NY: Extere Software. 7 p.

Rolletschek, H., A. Rolletschek, H. Kuhl, and J.-G. Kohl. 1999. Clone specificdifferences in a Phragmites australis stand. II. Seasonal development ofmorphological and physiological characteristis at the natural site and aftertransplantation. Aquat. Bot. 64:247–260.

Romesburg, H. C. 1984. Cluster Analysis for Researches. Belmont, CA: LifetimeLearning Publications. 344 p.

Russell, J. R., J. D. Fuller, M. Macaulay, B. G. Hatz, A. Jahoor, W. Powell, andR. Waugh. 1997. Direct comparison of levels of genetic variation amongbarley accessions detected by RFLPs, AFLPs, SSRs and RAPDs. Theor. Appl.Genet. 95:714–722.

Saltonstall, K. 2002. Cryptic invasion by a non-native genotype of the commonreed, Phragmites australis in North American. Proc. Nat. Acad. Sci. U. S. A.99:2445–2449.

Saltonstall, K. 2003a. Microsatellite variation within and among North Americanlineages of Phragmites australis. Mol. Ecol. 12:1689–1702.

Saltonstall, K. 2003b. A rapid method for identifying origin of North AmericanPhragmites populations using RFLP analysis. Wetlands 23:1043–1047.

Silander, J. A., Jr. 1985. Microevolution in clonal plants. Pages 107–152 inJ.B.C. Jackson, L. W. Buss, and R. E. Cook, eds. Population Biology andEvolution of Clonal Organisms. New Haven, CT, and London: YaleUniversity Press.

Stebbins, G. L. 1971. Chromosomal Evolution in Higher Plants. London:Edward Arnold. 216 p.

Swofford, D. L. and G. J. Olsen. 1990. Phylogeny reconstruction. Pages 411–501in D. M. Hillis and C. Moritz, eds. Molecular Systematics. Sunderland, MA:Sinauer Associates.

Weber, E. and B. Schmid. 1998. Latitudinal population differentiation in twospecies of Solidago (Asteraceae) introduced into Europe. Am. J. Bot.85:1110–1121.

Widen, B., N. Cronberg, and M. Widen. 1994. Genotypic diversity, molecularmarkers and spatial distribution of genes in clonal plants, a literature survey.Folia Geobot. Phytotax. Praha. 29:245–263.

Williams, J.G.K., A. R. Kubelik, K. J. Livak, J. A. Rafalski, and S. V. Tingey.1990. DNA polymorphisms amplified by arbitrary primers are useful asgenetic markers. Nucleic Acids Res. 18:6531–6535.

Zeidler, A., S. Scheneiders, C. Jung, A. E. Melchinger, and P. Dittrich. 1994. Theuse of DNA fingerprint in ecological studies of Phragmites australis (Cav.).Trin. ex Steudel. Bot. Acta 107:237–242.

Received October 30, 2010, and approved March 29, 2011.


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