Egypt. J. Plant Breed. 23(2):289– 307 (2019)

MAPPING QTL AND EPISTATIC EFFECTS FOR

POWDERY MILDEW, LEAF RUST AND NET BLOTCH

RESISTANCE IN BARLEY M.A. Sayed1, M. Abo Zaid2 and M.B. Ali1

1. Agronomy Dept., Fac. of Agri., Assiut University, Assiut 71526, Egypt

2. Wheat Diseases Research, Plant Pathology Res. Inst., ARC, Giza, Egypt

ABSTRACT Powdery mildew, leaf rust and net blotch are major biotic stresses that affect

both the quality and quantity of the yield of barley. The current research was conducted

at Sids Research Station during 2016/17 and 2017/18 growing seasons in Egypt. We

aimed to identify the quantitative trait loci (QTL) and digenic epistatic interactions

controlling the resistance to the aforementioned diseases in a doubled haploid

population (S42) of barley derived from the cross between the exotic accession of H.

vulgare ssp. spontaneum ‘’ISR42-8’’ and the German spring barley cultivar ‘’Scarlett’’

(H. vulgare ssp. vulgare). Results indicated that the wild parent is more resistant to the

investigated diseases than the cultivated barley; in addition, a favorable transgressive

segregation was detected for all investigated diseases. The QTL analysis showed that the

alleles of the marker bPb-9668 (4H, 145.02 cM) displayed favorable performance of

decreasing powdery mildew severity in the S42 population. In addition, three QTL were

mapped to 2H, 3H and 5H, which associated with the resistance of leaf rust. The alleles

of the three detected QTLs showed favorable performance of decreasing leaf rust

severity in the S42 population by values ranging between -33.54 and -17.73 %. The locus

QTS.S42.3H that located on chromosome 3H, exhibited both marker main effect and

marker × environment interaction. Furthermore, the exotic alleles of this locus

exhibited favorable performance of decreasing net blotch severity in the S42 population.

Moreover, the epistasis analysis revealed nine desirable pairs of epistatic effects

responsible for reducing the severity of the previous mentioned diseases in the S42

population. Our study highlighted that additive × additive epistasis was significant in the

inheritance and breeding for the resistance of the investigated barley diseases.

Key words: Barley, Blumeria graminis hordei, Puccinia hordei , Pyrenophora teres,

QTL analysis.

INTRODUCTION

Today, barley (Hordeum vulgare L.) is a very important cereal crop

worldwide. In this regard, it occupies the fourth rank after wheat, maize,

and rice (FAOSTAT 2018). Diseases that affect the foliage of the barley

plants such as powdery mildew (Blumeria graminis f. sp. hordei), leaf rust

(Puccinia hordei ) and net blotch (Pyrenophora teres Drechs) can severely

reduce the photosynthetic capacity of the plants and may produce toxins.

Furthermore, these diseases often result in yield reduction and excessive

grain protein for malting due to decreased grain size, number of grains per

spike and 1000-grain weight (Mathre 1997 and Ma et al 2004).

Developing disease resistant cultivars is a sustainable way to control

diseases (Mathre 1997 and Bajgain et al 2016). The exotic germplasm

including land races, and wild barley accessions may possess the resistance

to most economically crucial biotic and abiotic stresses (Ullrich et al 1995).

These germplasms can be used to develop new disease-resistant varieties in

290

barley breeding programs (Tanksley and Nelson 1996 and Gur and Zamir

2004). In this regard, molecular approaches will increase the economic

significance of breeding for diseases resistance (Parlevliet 1993). One of

the most vital cases for the introduction of favorable wild barley alleles into

elite barley cultivars is the mlo powdery mildew resistance (Von Korff et al

2005).

Numerous studies have extensively applied the QTL analysis in

barley and determined that the aforementioned diseases resistance genes are

present on various barley chromosomes (Ma et al 2004, Friesen et al 2006,

Abu-Qamar et al 2008, Hickey et al 2011, Liu et al 2011 and Li and Zhou

2011). The PI466197 is one of the powdery mildew resistance genes from

H. vulgare ssp. spontaneum, which was identified on the short arm of

chromosome 1H. The other gene for resistance was recognized on the short

arm of chromosome 2H (Teturova et al 2010). In addition, eleven

associated genes conferring resistance to the powdery mildew fungus

Blumeria graminis f.sp. hordei (Bgh) were detected by Spies et al (2012).

Rust-resistant QTL effective at seedling and adult plant stages have been

mapped on chromosome 6H (Marcel et al 2007 and Varshney et al 2007)

and, on the short arm of chromosome 5H by Hickey et al (2011) and Li and

Zahou (2011). The QTL analysis of the doubled haploid (DH) population

that originated from the cross between SM89010 and Q21861 identified

major position for net blotch resistance on chromosome 6H (Friesen et al

2006). This gene was mapped using the simple sequence repeat markers

and was found on a similar location (Cakir et al 2003). Our main objective

in the current study was to identify the beneficial and valuable exotic

alleles, which are vital in the expression of the resistance to powdery

mildew, leaf rust and net blotch in DH population of barley under Egypt

conditions using QTL mapping and epistasis analysis.

MATERIALS AND METHODS

Plant materials

An advanced backcross doubled-haploid mapping population (AB-

DH) containing 301 doubled haploid lines (DHs) derived from crossing

between the exotic accession of H. vulgare ssp. spontaneum ‘’ISR42-8’’

and the German spring barley cultivar ‘’Scarlett’’ (H. vulgare ssp. vulgare

L.) was used to detect QTLs controlling resistance for three diseases i.e.

powdery mildew, leaf rust and net blotch. The population was assigned as

S42 and utilized for QTL analysis and epistasis in current study. The elite

German cultivar Scarlett was used as the recurrent parent whereas the

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exotic one ISR42-8, which was utilized as the donor. Further details about

the development of S42 population and the proportion of exotic parent

genome can be found in von Korff et al (2004).

Molecular characterization of S42 population

A set of 371 doubled haploid lines was genotyped by using DNA

markers comprising 106 SSRs as per Von Korff et al (2004), 255 DArT

according to Sayed et al (2012) and ten gene-specific DNA markers as per

Wang et al (2010) to perform QTL analysis. The genetic map of current

population was drawn by MapChart ver.2.2 (Voorrips 2002).

Pathogen material

The S42 population and its parents were screened in Egypt against

three diseases i.e. powdery mildew (Blumeria graminis f. sp. hordei), leaf

rust ((Puccinia hordei) and net blotch at Sids Research Station during

2016/17 and 2017/2018 growing seasons. Net blotch is a seed-borne and a

foliar disease of barley (Douglas and Gordon 1985) caused by the fungus

Drechslera teres (Sacc.) Shoem. (Syn. Helminthosporium teres Sacc.), the

conidial state of Pyrenophora teres (Died) Drechslera. Each barley

genotype was sown in two rows of 1m length with two replicates using a

randomized complete blocks design. Recommended agricultural barley

practices were applied and the plots were surrounded by spreader area

planted with a mixture of highly susceptible barley varieties to spread

inoculum for infection rows to increase the disease pressure inoculum. For

field inoculation with leaf rust, the spreader plants were sprayed with a mist

of water and dusted with a mixture of urediniospores of the prevalent rust

races mixed with talcum powder at a rate of 1 (spores): 20 (talcum powder).

The inoculation of all barley plants was carried out at booting stage

according to the method of Tervet and Cassell (1951).

Diseases assessment

Diseases assessment of three barley diseases was covered out

through two growing seasons as follows: leaf rust assessment was

preformed when the susceptible check variety of barley expressed 50% rust

severity. The leaf rust severity (%) was recorded based on the modified

Cobb’s scale on a 0-100% scale (Peterson et al. 1948). The powdery

mildew was evaluated using a visual scoring scale with 9 classes. The first

class corresponds to an absence of symptoms, and the ninth indicates to an

attack similar to the ones observed on the susceptible control variety.

The reaction of barley plants to net blotch disease was recorded at

start of flowering stage (Larg 1954), according to double-digit scale 00-99.

292

The first digit gives the relative height of disease (Sarri and Prescott 1975)

whereas the second digit shows the disease severity as percentage of leaf

area affected in terms of 0-9, where: 0 = 0% severity, 2 = 20%, 3 = 30%

and so on (Eyal et al 1987).

Analysis of variance

To detect the variation among the tested DH lines and between

ISR42-8 and Scarlett for the investigated traits, the analysis of variance was

conducted using the Statistical Analysis System SAS software (SAS

Institute v. 9.2 2008), PROC GLM procedure. To meet the assumption

underlying the analysis of variance model, the data set of the three diseases

was transformed by calculating the square root of each value after adding 2.

Broad-sense heritability estimation

Heritability in broad-sense (H2) as percentage was estimated as H2=

[δ 2G/(δ2G + δ2

e/r)]×100, where δ2G and δ2

e were the estimates of genetic and

error variances, respectively, derived from the expected mean squares of

the separate analysis for each season. For estimation of broad-sense

heritability (H2) as percentage from combined analysis of variance, the H2

was computed as H2= [δ 2G / (δ2

G + δ 2G×Y/y + δ2

e/yr)]×100, where δ2G,

δ2G×Y and δ2

e were the estimates of genetic, genotype × years interaction

and error variances, respectively. Where, y and r were the number of years

and replications, respectively.

Relative performance of the wild accession RP[Hsp]

The relative performance of the wild parent was calculated using the

following equation:

RP[Hsp] = ([Hsp]- [Hv]) × 100/ [Hv]

where Hsp denotes LS-means of the homozygous wild genotype and Hv

symbolizes LS-means of the elite genotype.

QTL and epistasis analyses

The marker main effect (QTL), its genetic effect and the digenic

epistatic interactions among DNA marker pairs were examined using

stepwise regression approach using QTL mapper software version 1.60

(Wang et al 1999).

RESULTS AND DISCUSSION

Diseases performance and heritability estimates

The analysis of variance means and range between ISR42-8 and

Scarlett for powdery mildew, leaf rust and net blotch diseases compared to

their progeny are shown in Table (1).

293

Table 1. Mean squares, coefficient of determination (R2), heritability

estimates (H2), means and range of powdery mildew, leaf

rust and net blotch diseases for ISR42-8 and Scarlett the

compared to their S42 population.

Traits

LS-means ± SE Range

MS R2 H2

S42 population

ISR 42-8 Scarlett ISR 42-8 Scarlett LS-mean ±

SE Range

Powdery

mildew 1.41 ± 0.01 4.21 ± 0.75 1.41-1.41 3.46-6.48 15.73* 84.87 92.77 1.84 ± 0.14 1.41 - 7.21

Leaf rust 1.57 ± 0.14 7.69 ± 0.31 1.41 - 2.00 7.21 - 8.48 74.85** 98.68 99.55 2.95 ± 0.19 1.41 - 9.05

Net blotch 1.41 ± 0.01 7.70 ± 0.16 1.41-1.41 7.21 - 7.87 79.23** 99.79 99.93 3.18 ± 0.16 1.41 - 9.59

*, ** Indicate the significance level at 0.05 and 0.01, respectively.

Highly significant differences between both parents for the

resistance to the three studied diseases were observed in this sudy. The wild

accession ISR 42-8 showed low values of mean disease severity and

recorded 1.41, 1.57 and 1.41 for powdery mildew, leaf rust and net blotch,

respectively. On the other hand, the cultivated parent Scarlett showed high

mean values of the disease severity and recorded 4.21, 7.69 and 7.70,

respectively. This result indicates that the wild parent is more resistant to

the investigated diseases than the cultivated one. Cherif et al (2008)

reported that the mean value of the DH lines was significantly higher than

the mid-parent for net blotch disease studied in three environments.

High estimates of both coefficient of determination and heritability

in broad sense that calculated from both parents were detected for all

investigated diseases. In comparison of their parents, the S42 population

showed wide range of the disease severity of the three diseases. Moreover,

based on extreme values of the S42 population, favorable transgressive

segregation was detected for all investigated diseases, since the DH lines

showing better performance than Scarlett (Table 1 and Figures 1, 2 and 3).

For the S42 population, the analysis of variance showed highly significant

differences among DH lines for all investigated diseases (Table 2),

indicating the suitability of this population to the QTL analysis. Both

seasons differed significantly only in powdery mildew disease case and

there was no interaction between years and DH lines for all investigated

diseases. Furthermore, net blotch disease showed the minimum values of

the coefficient of variation in both seasons and overall ones, while powdery

mildew, exhibited the maximum values of the coefficient of variation.

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Table 2. Separate and combined analysis of variance, coefficient of

determination (R2) and heritability estimates (H2) of powdery

mildew, leaf rust and net blotch diseases in the S42 barley

population.

Source DF

Powdery mildew Leaf rust Net blotch

MS C.V. R2 H2 MS C.V. R2 H2 MS C.V. R2 H2

Separate analysis of 2016/2017 season

Rep 1 1.84* 38.12 83.04 79.52 1.74 24.67 90.04 88.93 4.30** 22.81 95.84 95.65

Lines 300 2.21** 4.84** 11.96**

Error 300 0.45 0.54 0.52

Separate analysis of 2017/2018 season

Rep 1 0.13 20.98 95.97 95.81 0.03 28.06 88.05 86.43 0.03 17.44 97.54 97.48

Lines 300 3.87** 4.99** 12.34**

Error 300 0.16 0.68 0.31

Combined analysis

Year 1 7.38* 30.09 90.86 93.46 0.32 26.40 89.02 92.74 0.40 20.27 96.69 97.88

Year(Rep) 2 0.99 0.89 2.17

Lines 300 5.77** 9.57** 24.14**

Year×Lines 300 0.34 0.65 0.47

Error 600 0.31 0.61 0.42

*, ** Indicate the significance level at 0.05 and 0.01, respectively.

295

0 2 4 6 8 10

0

50

100

150

200First season

Scarlett

ISR 42-8

Powdery mildew scalePowdery mildew scale

2 4 6 8

0

50

100

150

200

250

300

Num

ber

of D

H l

ines

Second season

Scarlett

ISR 42-8

Num

ber

of D

H l

ines

Fig. 1. Frequency distribution of population S42 for powdery mildew

severity scale at the first and second season. The vertical lines

represent mean trait value of Scarlett and ISR42-8.

0 2 4 6 8 10

0

20

40

60

80

100

First season

Scarlett

ISR 42-8

Num

ber

of D

H li

nes

0 2 4 6 8 10

0

20

40

60

80

100

Second season

Scarlett

ISR 42-8

Num

ber

of D

H li

nes

Leaf rust scale Leaf rust scale

Fig. 2. Frequency distribution of population S42 for leaf rust severity

scale at the first and second season. The vertical lines represent

mean trait value of Scarlett and ISR42-8.

296

0 2 4 6 8 10

0

50

100

150

200 First season

Scarlett

ISR 42-8

Num

ber of D

H lines

Num

ber of D

H lines

Net blotch scale Net blotch scale

0 2 4 6 8 10

0

50

100

150

200 Second season

ScarlettISR 42-8

Fig. 3. Frequency distribution of population S42 for net blotch severity

scale at the first and second season. The vertical lines represent mean

trait value of Scarlett and ISR42-8.

In contrast, net blotch disease exhibited high estimates of broad

sense heritability in both seasons and as combined overall seasons (Table

297

2). The frequency distribution of the S42 population for the investigated

diseases in both seasons are shown in Figures 1, 2 and 3. These data

revealed skewed distribution for the three diseases. The large amount of

variation among barley DH lines in their reactions for the disease has also

observed by several researchers such as Von Korff et al (2005) and Cherif

et al (2008). Moreover, the transgressive segregation for both resistance

and susceptibility of the three diseases in separate and combined

environments were reported in several researches in barley (Von Korff et al

2005 and Hickey et al 2011).

Detection of QTL

Altogether, nine putative QTLs and marker × environment

interaction were detected for the investigated diseases in current study.

These QTL were distributed as four QTL detected in the first season and

five QTL were detected for the second season. As main effect overall

seasons, seven QTL were identified for the three diseases (Table 3 and

Figure 4). Among these loci, two QTL were detected for powdery mildew

and mapped on chromosomes 1H and 4H. The locus, QPM.S42.1H

exhibited significant marker main effects and marker × environment

interaction. Based on the relative performance of the exotic allele (RP[Hsp]),

the alleles of the marker bPb-5198 exhibited unfavorable performance of

increasing powdery mildew severity in the S42 population by values 21.88

and 31.73% in the first and second season, respectively. Whereas the

marker locus QPM.S42.4H showed marker × environment interaction and

the alleles of the marker bPb-9668 (4H, 145.02 cM) exhibited favorable

performance of decreasing powdery mildew severity in the S42 population

by values -1.37%. Von Korff et al (2005) detected nine putative QTLs for

powdery mildew covered the whole genome of the same population used in

current study. They found that six QTL out of nine, the exotic parent

contributed the favorable allele to the powdery mildew. Our results are in

accordance with those obtained by Von Korff et al (2005), since the marker

locus bPb-9668 (4H, 145.02 cM) is very near to the DNA region which

detected by Von Korff et al (2005) and also corresponds to the Mlg locus.

Furthermore, Wei et al (1999) identified a race-specific locus Mla on

choromosome 1H encodes 32 different Bgh-resistant genes. Smilar results

were obtained by Teturova et al (2010).

298

Table 3. Detected QTLs for powdery mildew, leaf rust and net blotch

diseases in the S42 barley population at two seasons.

1)QTL 2)Season 3)Marker 4)Ch 5)Range 6)F

Value 7)Prob.

8)R2

(%) 9)Effect

10)Hv

effect

11)Hsp

effect 12)RP[Hsp] 13)Additive

Powdery mildew

QPM.S42.1

H 1 bPb-5198 1H

116.46-

123.07 8.94** < 0.003 2.83 M, M×E 1.68 2.05 21.88 0.184

QPM.S42.4

H 1 bPb-9668 4H

145.02-

145.10 7.2** < 0.007 2.28 M×E 1.91 1.88 -1.37 -0.013

QPM.S42.1

H 2 bPb-5198 1H

116.46-

123.07 12.25** < 0.0005 3.93 M, M×E 1.79 2.36 31.73 0.284

Leaf rust

QLR.S42.2

H 2 bPb-1815 2H

146.58-

146.63 9.99** < 0.0017 3.17 M, M×E 3.96 2.82 -28.82 -0.571

QLR.S42.3

H 2 bPb-8419 3H 153.55 8.08** < 0.0048 2.56 M, M×E 3.42 2.81 -17.73 -0.303

QLR.S42.5

H 1 bPb-0837 5H

18.03-

20.0 8.27** < 0.0044 2.69 M, M×E 3.05 2.03 -33.54 -0.511

Net blotch

QNB.S42.3

H 1 MGB410 3H 65-66 6.67* < 0.0104 2.18 M, M×E 4.40 3.06 -30.50 -0.670

QNB.S42.3

H 2 MGB410 3H 65-66 8.97** < 0.0039 2.91 M, M×E 4.43 3.09 -30.26 -0.671

QNB.S42.1

H 2 bPb-2240 1H

123.08 –

125.0 4.19* < 0.029 1.15 M×E 2.99 3.68 22.93 0.343

1) Description of quantitative trait locus. 2) Season. 3) Linked DNA marker

revealing strongest F-value. 4) Chromosome. 5) The QTL region associated to

the significant DNA markers in centiMorgan positions. 6) F-value. 7)

Probability of F test at P < 0.01. 8) Genetic variance explained by DNA

markers. 9) M Marker main effect, M × E marker × environment interaction.

10) Trait value of homozygous cultivated genotypes (Hv). 11) Trait value of

homozygous exotic genotypes (Hsp). 12) Relative performance of the

homozygous exotic allele, RP [Hsp] and 13) The additive effect is half the

difference between the phenotypic means of the homozygous elite and exotic

marker genotypes. *, ** Indicate the significance level at 0.05 and 0.01,

respectively

299

MGB4020.0bPb-28624.3bPb-63184.8bPb-04057.2bPb-62387.5bPb-377610.5bPb-960811.5bPb-713711.7bPb-131813.1GMS2114.0S5370718.0GBM100719.0bPb-730619.1bPb-441536.6Mla1238.5GBM104239.0bPb-321740.5bPb-048255.4bPb-160455.8bPb-971758.7bPb-453160.2bPb-360562.2HVALAAT62.5HVM2062.6Bmag21162.7MGB32562.9Bmag14963.2Bmag10563.5HVGLUEND64.0bPb-529064.9bPb-533467.9bPb-533976.8Bmac3280.0bPb-760982.1bPb-789986.3bPb-691194.9bPb-121395.0bPb-136695.1bPb-4515106.2HvFT3115.0HVABAIP116.0bPb-5014bPb-5198

116.5

bPb-2240123.1GBM1061125.0bPb-1882133.1bPb-6200133.7GBMS12134.0bPb-1940135.1bPb-6502139.6bPb-8112140.9bPb-0589141.2bPb-5201141.3bPb-0699144.2bPb-1487147.3GBMS143162.0

QP

M.S

42

.1H

QN

B.S

42

.1H

1H

x0.0bPb-42853.5bPb-51915.0bPb-96815.3bPb-64667.6bPb-975714.4bPb-8399bPb-0003

25.7

bPb-612826.2HVM3626.5GBM103527.0bPb-829227.1bPb-305030.2bPb-482135.8bPb-803839.0PpdH141.1GBM105242.0bPb-426144.8bPb-790647.4bPb-223060.4MGB39164.0HvGOGAT65.0HvFT466.0bPb-222567.3bPb-107270.0bPb-305670.8bPb-877977.4EBmac68480.0GMS381.0bPb-608881.7bPb-404082.1bPb-975482.8bPb-089087.0HvNAM290.0HVTUB92.0Bmag38197.0Bmag12598.0bPb-814398.2bPb-7991101.3bPb-1926102.1bPb-3563102.4bPb-4577108.7bPb-8464138.2bPb-7816139.0bPb-4768139.6bPb-1066139.8GBM1016140.0HVM54143.0EBmac415144.0HvCNX1145.0bPb-1815bPb-4092

146.6

bPb-8255149.4MGB334150.0bPb-5558150.1bPb-0299157.1bPb-3102160.4bPb-8698161.1bPb-7723163.3

QL

R.S

42

.2H

2H

x0.0bPb-44721.5bPb-95836.0bPb-36897.6bPb-30259.9bPb-994510.2bPb-107720.0HVLTPPB25.0EBmac70530.0HVITR135.0bPb-798950.4bPb-793851.4bPb-299351.6bPb-727353.2bPb-0158bPb-9746

54.8

bPb-634755.6HvGI63.0HvFT264.0MGB41065.0Bmag60366.0bPb-204066.5bPb-5771bPb-0094

69.3

bPb-828369.6bPb-501269.8bPb-832170.4HVM3383.0bPb-035384.4bPb-168187.8GMS116100.0HVM60100.1GBM1043100.7bPb-3278100.8bPb-4616105.9bPb-4209111.7bPb-1579115.5bPb-9110118.7bPb-1609140.3bPb-9111141.9bPb-4564142.8bPb-1481145.4bPb-5129146.8bPb-2420147.0bPb-6228bPb-8557

147.9

bPb-7827148.3bPb-0789148.8bPb-2888149.8HV13GEIII150.0HVM62151.0MGB358152.0bPb-8419153.5bPb-7164158.0bPb-1411160.2bPb-0361165.5bPb-5864170.7bPb-4628175.2Bmac29176.0bPb-7247178.5bPb-8962178.6bPb-7724179.5bPb-0136181.3

QL

R.S

42

.3H

QN

B.S

42

.3H

3H

x0.0

bPb-930412.7bPb-693813.0HVM4014.0bPb-146914.6bPb-469917.3HVOLE21.0HVB23D25.0HVKNOX331.0

HVPAZXG44.0

HVM1355.0GMS8957.0bPb-1408bPb-8437

60.0

bPb-664060.5bPb-351267.9bPb-421669.7bPb-548072.2bPb-798772.3EBmac77580.0bPb-801386.7bPb-870193.6MGB39695.0bPb-373996.3bPb-771996.8

bPb-9859123.2TACMD125.0Mlo127.5EBmac701130.0EBmac635131.0EBmac679132.0EBmac788138.0GBM1015140.0VrnH2140.2HVJASIP141.0HVM67141.1bPb-9820142.1bPb-9668145.0bPb-5265145.1HDAMYB146.0bPb-3717148.6

QP

M.S

42

.4H

4H

x0.0

bPb-083718.0BMS220.0Bmac16324.0

MGB38433.0bPb-932734.7bPb-636336.1

bPb-413543.5

bPb-728758.4Bmag33765.0Bmag35768.0bPb-961870.7bPb-776371.0bPb-785273.6bPb-667681.4bPb-237882.9bPb-2497bPb-4721

84.2

Bmag22387.0bPb-6967MGB338

95.0

bPb-124199.9bPb-5596101.3bPb-6126106.9

VrnH1125.1GMS61126.0bPb-4758126.5bPb-0071126.8bPb-3910133.5bPb-3945136.2bPb-5845139.0MGB318150.0AF043094A156.0bPb-6367157.4bPb-5238159.4bPb-0171159.9Bmag222162.0MGB357165.0bPb-3138166.6bPb-1719173.7bPb-0799174.0bPb-8754183.0bPb-1217184.4bPb-9660187.0

QL

R.S

42

.5H

5H

Fig. 4. Genetic map of the detected QTL for powdery mildew, leaf rust

and net blotch in barley. Linkage map was drawn using

MapChart ver.2.2 where the markers and genetic positions are

presented on right and left of the chromosome, respectively.

300

For leaf rust disease, the QTL analysis identified three QTL and

mapped to 2H, 3H and 5H. All detected QTL showed marker main effect

and marker × environment interaction. The alleles of the three detected

QTL exhibited favorable performance of decreasing leaf rust severity in the

S42 population by values ranging between -33.54and -17.73%. These QTLs

showed negative additive effects (Table 3). The strongest QTL was QLR.

S42. 2H (146.58 cM) and explained 3.17% of the genetic variance. This

DNA region maps to the same bin as a gene, which encodes chalcone

synthase, the chalcone synthase is responsible of the defense response

genes and was accumulated in barley leaves upon inoculation with powdery

mildew (Christensen et al 1998 and Karakousis et al 2003). Von Korff et al

(2005) identified six putative QTL for leaf rust resistance in the S42

population. One of them (EBmac415) was located on chromosome 2H (146

cM) and led to reduce the severity of leaf rust in the S42 population.

Furthermore, the detected QTL QLR.S42.3H which located on 3H (153.55

cM) was corresponding to the marker HVM62 (3H, 155 cM) which was

identified by Vvon Korff et al (2005). Several race-specific resistance

genes for leaf rust were reported in barley (Franckowiak et al 1997), some

of these genes were identified in H. vulgare ssp. Spontaneum. Many studies

detected resistance genes to leaf rust in barley on chromosomes 2H and 3H

such as Feuerstein et al (1990), who detected two resistance genes, Rph10

and Rph11, on chromosomes 3H and 6H, and Ivandic et al (1998) who

mapped Rph16 at the centromeric region of 2H. Hickey et al (2011)

detected genomic regions associated to leaf rust resistance in barley using

DArT markers. Remarkably, one of the important result of current study

that the marker locus bPb-0837 which was mapped on 5H was the same

marker which was detected by Hickey et al (2011) for leaf rust resistance.

Sandhu et al (2016) characterized some international barley nurseries in

Australia for leaf rust resistance and detected the resistance genes.

Barley net form net blotch is a significant foliar malady of barley

(Ilyas et al 2014). Two QTL were detected for net blotch disease (Table 3).

The locus QTS.S42.3H that located on chromosome 3H, showed marker

main effect and marker × environment interaction. In addition, the exotic

alleles of this locus exhibited favorable performance of decreasing net

blotch severity in the S42 population by values -30.50 and -30.26% in the

first and second season, respectively. The other locus, QTS.S42.1H, which

was mapped on chromosome 1H showed marker × environment interaction.

The exotic alleles of this locus exhibited unfavorable performance of

301

increasing net blotch severity in the S42 population by value of 22.93%. In

addition, the exotic alleles explain small percentage (1.15%) of the genetic

variance. Tajinder et al (2012) identified one spot blotch QTL on

chromosome 1H. In addition, the chromosome 6H has been reported to

contain multiple independent resistance genes against this disease (Friesen

et al 2006, Abu Qamar et al 2008 and Liu et al 2012).

Detection of Epistasis

Using the doubled haploid lines can provide valuable information

on additive and epistatic interaction effects that are maximized, since all

polymorphisms can be explained by dominant and co-dominant allele

expression (Choo et al 1985). Altogether, 18 pairs of epistatic QTLs as

additive-by-additive effects were identified for three investigated diseases

in both seasons in the S42 population (Table 4). Among the QTL detected,

only the marker locus bPb-2240 (1H) was involved in epistatic interaction

effects. The result indicates that several loci that involved in the epistatic

interactions may not have significant effects for the investigated diseases

and may affect the trait expression by epistatic interactions with other loci.

Results revealed seven pairs of epistatic QTLs (one for the first

season and six for the second season) were associated significantly with

powdery mildew and covered the whole genome except chromosome 5H.

Among these positions, two pairs of epistatic effects reduced powdery

mildew severity in the S42 population. The most desirable pair of epistatic

QTL for reducing powdery mildew severity was (bPb-3278* bPb-0432)

and mapped to chromosomes 3H (100.76 cM) and 6H (91.99 cM) and had

the highest F value and accounted for 5.21% of genetic variation (Table 4).

At this locus, the BC2DH lines carrying the Hsp/Hsp genotype were

resistant to the powdery mildew with an average of 1.63 of disease scale

compared to the other the allelic combinations. Using scaling test, Fazeli et

al (2013) indicated the importance role of additive × additive effect in

controlling of powdery mildew resistance in barley.

302

Table 4. Estimation of LS-means of 17 pairs of digenic interactions and

epistatic effects (additive × additive) for powdery mildew,

leaf rust and net blotch diseases in each season.

Marker 1 Marker 2 F Value Prob. F R2

LS means of digenic interactions

Name Chr. Pos. Name Chr. Pos. Hv/Hv Hv/Hsp Hsp/Hv Hsp/Hsp

Powdery mildew, season 1

bPb-1366 1H 95.08 bPb-3512 4H 67.92 10.25** <

0.0015 3.15 1.68 1.76 1.72 1.79

Powdery mildew, season 2

MGB391 2H 64.00 bPb-7247 3H 178.50 9.57** <

0.0022 2.91 1.83 2.01 2.01 2.08

bPb-7609 1H 82.15 HVPAZXG 4H 44.00 9.67** <

0.0021 2.59 1.69 2.22 1.76 2.29

HvCO1 7H 82.00 bPb-8690 7H 87.39 15.72** <

0.0001 4.21 1.86 1.96 1.87 1.97

S53707 1H 18.00 HVM60 3H 100.10 8.41** < 0.004 2.25 1.93 1.92 1.93 1.92

bPb-3278 3H 100.76 bPb-0432 6H 91.99 19.47** <

0.0001 5.21 2.04 1.76 1.92 1.63

bPb-5201 1H 141.29 bPb-3895 6H 98.71 14.56** <

0.0002 3.9 1.84 1.89 2.08 2.12

Leaf rust, season 1

bPb-3217 1H 40.53 HVPAZXG 4H 44.00 9.73** < 0.002 2.74 3.17 3.33 2.49 2.65

bPb-2230 2H 60.45 bPb-4216 4H 69.66 5.02* < 0.025 1.42 3.04 2.73 2.67 2.36

bPb-1815 2H 146.58 HVPAZXG 4H 44.00 22.00** <

0.0001 6.2 2.81 3.05 3.35 3.63

bPb-9681 2H 5.27 AF043094A 5H 156.00 10.48** <

0.0014 2.95 2.99 2.69 2.91 2.61

HVKNOX3 4H 31.00 HVCHI26A 7H 159.20 16.17** <

0.0001 4.56 3.00 2.48 3.01 2.48

Leaf rust, season 2

bPb-9681 2H 5.27 AF043094A 5H 156.00 12.09** <

0.0012 2.97 3.01 2.47 2.66 2.32

Net blotch, season 1

bPb-2240 1H 123.09 bPb-9304 4H 12.67 12.74** <

0.0004 3.94 3.13 4.09 3.44 4.40

Bmag357 5H 68.00 bPb-6967 5H 94.96 9.48** <

0.0023 2.93 3.10 3.07 3.42 3.39

Net blotch, season 2

bPb-1604 1H 55.77 bPb-9820 4H 142.09 6.89* <

0.0107 2.06 3.20 4.26 3.49 4.55

bPb-6727 6H 134.08 bPb-6706 7H 58.17 10.64** <

0.0017 3.18 3.73 2.78 2.60 1.65

EBmac635 4H 131.00 HvCO1 7H 82.00 8.41** <

0.0051 2.52 3.39 2.98 3.11 2.70

*, ** indicate the significance level at 0.05 and 0.01, respectively to declare the

putative epistatic QTL positions.

303

For leaf rust, the analysis revealed six pairs of epistatic QTL (five

for the first season and one for the second season) were associated

significantly with leaf rust disease. These QTL were located on

chromosomes 1H, 2H, 4H, 5H and 7H. Among these locations, five pairs of

epistatic effects reduced leaf rust severity in the S42 population. The most

desirable pair of epistatic QTL for reducing leaf rust severity was

(HVKNOX3 × HVCHI26A) located on chromosomes 4H (31.00 cM) and

7H (159.20 cM) and had the highest F value and accounted for 4.56% of

genetic variation (Table 4). At this locus, the BC2DH lines carrying the

Hsp/Hsp genotype were resistant to the leaf rust with an average of 2.48 of

disease scale compared to the other the allelic combinations. In addition,

the epistatic pair bPb-9681 × AF043094A was observed in both seasons

and led to reducing leaf rust severity and had high F value and R2. Qi et al

(1998) detected an additive effect QTL and a significant interaction

between two QTL, i.e. Rphq1 and Rphq2.

For net blotch, the analysis revealed five pairs of epistatic QTL (two

for the first season and three for the second season) were associated

significantly with net blotch disease. These QTL were mapped on

chromosomes 1H, 4H, 5H, 6H and 7H. Among these loci, two pairs of

epistatic effects reduced net blotch severity in the S42 population. The most

favorable pair of epistatic QTL for reducing leaf rust severity was (bPb-

6727 × bPb-6706) and located on chromosomes 6H (134.08 cM) and 7H

(58.17 cM) and had the highest F value and accounted for 3.18% of genetic

variation (Table 4). At this locus, the BC2DH lines carrying the Hsp/Hsp

genotype were resistant to the net blotch with an average of 1.65 of disease

scale compared to the other the allelic combinations. Cherif et al (2008)

concluded that net blotch resistance is quantitative and additive × additive

epistasic effect plays an important in the inheritance of net blotch.

component of this resistance.

In conclusion, our results revealed that five favorable QTLs were

responsible of decreasing powdery mildew, leaf rust and net blotch severity

in the S42 population due to the presence of the exotic alleles of the wild

parent ISR 42-8. These multiple disease resistance regions were considered

as prime candidates for further investigation and validation of their broad

resistance. Further, the epistasis analysis revealed nine desirable pairs of

epistatic effects of reducing the severity of the abovementioned diseases in

the S42 population. Since, the DHs lines carrying the Hsp/Hsp genotype

were resistant to these diseases. Thus, the study highlighted that additive ×

304

additive epistasis was significant in the inheritance and breeding for the

resistance of the investigated diseases in barley.

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التفوقي للمقاومة ألمراض البياض الدقيقي, تحليل مواقع الصفات الكمية والتأثير صدأ األوراق و التبقع الشبكي في الشعير

1, محمد بدري محمد علي2, محمد أبوزيد1محمد عبدالعزيز عبدالحليم سيد قسم المحاصيل، كلية الزراعة، جامعة أسيوط . 1

بات، مركز البحوث الزراعية، الجيزةراض القمح، معهد أمراض النأم قسم. 2

, والتبقع الشبكي من اإلجهادات الحيوية الرئيسية التي تؤثر على يعتبر البياض الدقيقي, وصدأ األوراقو 2016/17جودة وكمية محصول الشعير. تم إجراء هذا البحث في محطة بحوث سدس خالل موسمي

( ودراسة التأثيرات التفاعلية QTLان الهدف من البحث, تحديد مواقع الصفات الكمّية )في مصر. ك 2017/18في المقاومة لألمراض الثالث في عشيرة من الشعير )مضاعفة العدد الكروموسومي األحادي(. المضيفة المتحكمة

وصنف ’’ H. vulgare ssp. spontaneum’’ ISR42-8هذه العشيرة ناتجة من التهجين بين األب البري كثر مقاومة النتائج إلى أن األب البري أ أشارت H. Vulgare ssp vulgare’’ Scarlett’’.شعير ألماني منزرع

لألمراض الثالثة من الشعير المنزرع. كما لوحظ وجود انعزاالت فائقة الحدود لمقاومة األمراض المذكورة. كشف أظهرت سنتيمورجانH4 (145.02 )روموسوم الك علي bPb-9668 أن أليالت الواسم الوراثي QTLتحليل

. باإلضافة إلى ذلك , تم تحديد ثالثة S42عشيرة أداء مرغوبًا لخفض شدة االصابة بمرض البياض الدقيقي في الQTL مواقع وراثية على الكروموسوماتH2 ,H3 ,H5 والتي ترتبط بمقاومة صدأ األوراق. أظهرت أليالت الـ ,QTL لتقليل شدة االصابة بصدأ األوراق في العشيرة ة أداًء مرغوبًا الثالثة المكتشفS42 بقيم تراوحت بين-

كال من التأثير الرئيسي H3الموجود على الكروموسوم QTS.S42.3Hكما أظهر الموقع ٪.17.73-و 33.54والتأثير التفاعلي بين الواسم والبيئة. عالوة على ذلك, أظهرت األليالت البرية لهذا الموقع أداًء مواتيًا لخفض شدة

عالت األليلية المرغوب فيها و تسعة أزواج من التفا Epistasis. كشف تحليل S42التبقع الشبكي في العشيرة ألصابة باألمراض المذكورة أعاله في العشيرة موضع الدراسة. تسلط هذه الدراسة المسؤولة عن الحد من شدة ا

المضيف في الوراثة والتربية للمقاومة لألمراض الثالثة التي تم دراستها في -الضوء على أهمية التفاعل المضيف هذه التجربة في الشعير.

(2019) 307 -289( : 2)23لمجلة المصرية لتربية النبات ا