Allelic Relationship between <i>Lr</i>9 and the Leaf Rust Resistance Gene in Kharchia Local Mutant of Wheat

doi:10.4236/ajps.2011.25083

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American Journal of Plant Sciences, 2011, 2, 688-691

doi:10.4236/ajps.2011.25083 Published Online November 2011 (http://www.SciRP.org/journal/ajps)

Allelic Relationship between Lr9 and the Leaf Rust

Resistance Gene in Kharchia Local Mutant of

Wheat

Navjot Kaur Dhillon1*, Harcharan Singh Dhaliwal2

1Department of Plant Breeding and Genetics, Punjab Agricultural University, Ludhiana, India; 2 Akal School of Biotechnology, Baru

Sahib, Himachal Pradesh, India.

Email: *dhillon.na[email protected]

Received August 9th, 2011; revised September 30th, 2011; accepted October 30th, 2011.

ABSTRACT

To confirm allelic relationship between Lr9 and the leaf rust resistance gene in KLM4-3B, genetics of resistance was

studied using crosses (WL711 + Lr9) × WL711 and (WL711 + LrKLM4-3B) × WL711. The F2 populations in cross

(WL711 + Lr9) × WL711 and (WL711 + LrKLM4-3B) × WL711 segregated in ratio of 3:1 for disease reaction at seed-

ling stage against pathotype 77-5 of leaf rust. This suggests that rust resistance in these stocks are under the control of

single dominant genes. Further, to study allelic relationship between Lr9 and LrKLM4-3B, F2 population of the cross

(WL711 + LrKLM4-3B) × (WL711 + Lr9) was studied. A segregation ratio of 15:1 implies that the two genes Lr9 and

LrKLM4-3B are non-allelic genes.

Keywords: Lr9, Isogenic Lines, Non-Allelic Genes

1. Introduction

It is imperative to stabilize the wheat production by redu-

cing the losses due to various diseases including leaf rust,

stem rust, yellow rust, Karnal bunt etc. Among the dis-

eases, leaf rust caused by Puccinia recondita Roberage

ex. Desmaz f.sp. tritici is one of the most important and

devastating foliar diseases of wheat which cause signifi-

cant yield losses all over the world [1-8]. In all regions in

which wheat is grown, rusts have caused periodic severe

epidemics [9]. The rust accelerates foliage senescence re-

ducing cumulative light interception of the crop which

leads to reduced dry matter production [10]. Breeding for

resistance against leaf rust is an economical, efficient and

environmentally safe control measure to reduce these

losses [11]. Development of disease resistant varieties is

one of the most economical methods of control of dis-

eases like leaf rust. However, growing of rust resistant

varieties having single gene for resistance results in rapid

evolution of virulent biotypes of the pathogen, thereby

making the resistance gene ineffective and the variety

susceptible to rust. One of the ways to develop varieties

with durable rust resistance is to pyramid the genes for

resistance in a single variety [12]. It is difficult to pyra-

mid two or more disease resistance genes through con-

ventional means, particularly where the resistance genes

in question are effective against all the prevalent patho-

types. However, recent advances in molecular biology

have made it possible to pyramid several genes in single

line using marker assisted selection (MAS). Tagging of

genes with molecular markers is pre-requisite for MAS

[13].

A number of rust resistance genes, including those for

leaf rust resistance, have been transferred from wild rela-

tives of wheat into cultivated wheats [14,15]. In India,

from the analyses of 2630 samples collected from 17

states, one union territory and Nepal from 2005 to 2008,

31 races were identified among which eight were new

[16]. Most of these could not, however be exploited com-

mercially because of extensive linkage drag. One of the

leaf rust resistance genes, Lr9 transferred from Aegilops

umbellulata [17] and located on chromosome 6BL, has

no undesirable effect associated with it [18]. This gene is

effective against all the races of leaf rust currently pre-

valent in Northern India. Similarly, another leaf rust resi-

stance gene identified in (Kharchia local mutant KLM4-

3B) is also effective against all the prevalent leaf rust

pathotypes in Northern India. In the absence of virulence,

capable of differentiating Lr9 and KLM4-3B in the In-

Allelic Relationship between Lr9 and the Leaf Rust Resistance Gene in Kharchia Local Mutant of Wheat689

dian subcontinent KLM4-3B has been suspected to be

Lr9 rather than an induced mutant of Kharchia local [19].

High density molecular maps have been constructed in

several crops including rice, maize, tomato and Triticum

[20,21] and a number of genes of economic importance

have been tagged with series of molecular markers [22,

23]. Molecular markers closely linked to the genes for

rust resistance can be used not only for establishing alle-

lic relationships among resistant sources but also for their

pyramiding using marker assisted selection (MAS).

2. Material and Method

2.1. Disease Reaction Studies

Single spore culture of P recondita f.sp. tritici variants

77-5 (maintained on Agra local) were used for identifica-

tion of F2 seedling resistance genes.

2.2. Raising of Seedlings

Seeds of the parents (WL 711, KLM4-3B and Thatcher +

Lr9) were sown along with F2 populations of the three

crosses in bread boxes containing a mixture of farmyard

manure and sandy loam soil in equal proportions. Agra

local was also sown as susceptible check. The seedlings

were raised in glass house maintained at a temperature of

25˚C ± 1˚C. Relative humidity above 80 per cent was

maintained by using desert cooler. The bread boxes were

watered every day to maintain vigour of the seedlings.

First leaf of the seven day old seedling was inoculated

with homogeneous mixture of appropriate rust culture

and talc, keeping inoculum density of 6 - 10 urediospores

per microscopic field at a magnification of 100x under

light microscope. After inoculation the seedlings were

incubated at 100 percent relative humidity for 16 hours.

These seedlings were then transferred to growth cham-

bers.

2.3. Scoring the Infection Types

Fourteen days after inoculation, the infection type on the

seedlings was scored using a modification of the scale

given by Stakman [24]. The seedlings showing .infection

type 0, 1, 2 and X were classified as resistant, whereas

those with infection types 3 to 4 were classified as sus-

ceptible.

Further, disease reaction of F2 population of three dif-

ferent crosses at adult stage were scored by using Modi-

fied Cobb Scale by Peterson [25].

2.4. Statistical Analysis

Simple Chi-square (χ2) test was applied to fit appropriate

genetic ratios in F2 generation obtained from the three

crosses (WL 711 + Lr9) × WL 711, (WL 711 + LrKLM4-

3B) × WL 711 and (WL 711 + Lr9) × (WL 711 +

LrKLM4-3B). Chi-square value was calculated using the

following formula:



1d.f .







where,

n = Number of phenotypic classes.

d.f. = Degree of freedom.

O = Number of observed plants in a phenotypic class.

E = Number of expected plants in a phenotypic class.

3. Results and Discussion

In the present investigation, to confirm the allelic rela-

tionship between the leaf rust genes Lr9 and the resistant

gene in KLM4-3B, F2 generations of three crosses (WL

711 + Lr9) × WL 711, (WL 711 + LrKLM4-3B) × WL

711 and (WL 711 + Lr9) × (WL 711 + LrKLM4-3B)

were studied. The results pertaining to these studies are

presented here.

Results of the cross of isogenic line of the leaf rust re-

sistant gene of KLM4-3B (LrKLM4-3B) with the recur-

rent parent, WL711 are presented in Tables 1 and 2. Out

of 122 F2 plants, 86 were resistant and 36 susceptible to

leaf rust pathotype 77-5. The infection type observed on

resistant plants were 0, 0, 1, 1+, 2, 2+. The segregation of

F2 plants showed a good fit to 3:1 ratio (χ2 = 1.32). This

indicated that the Lr KLM4-3B is dominant.

In the second cross of isogenic lines of Lr9 with the

recurrent parent, out of 126 F2 plants 89 plants were re-

sistant and 37 were susceptible to the leaf rust pathotype

77-5 [Table 1]. The ratio of resistant to susceptible

plants did not differ significantly from 3:1 (χ2 = 1.28).

This indicated that Lr9 also behaves as dominant gene to

pathotypes 77-5.

In the cross between isogenic lines carrying Lr9 and

LrKLM4-3B, out of 101 F2 plants, 91 showed resistant

reaction and 10 were susceptible to pathotype 77-5. This

did not differ significantly from 15 resistant: 1 suscepti-

ble ratio (Table 1). This suggested that the two leaf rust

resistant genes, Lr9 and LrKLM4-3B, are non-allelic.

The earlier studies have also shown that these two leaf

rust genes are non-allelic [19]. Lr9 is an alien gene on

chromosome 6BL translocation from Aegilops umbellata

[17], whereas LrKLM4-3B was identified to be resistant

to leaf rust [26]. Preliminary work carried out at the

School of Biotechnology, Punjab Agricultural University

has shown that the LrKLM4-3B is not a mutant gene as

claimed earlier [26] but is associated with translocation

involving chromosome 2BL (Dhaliwal and Harjit Singh,

Pers.Commu.). These observations further support that

these two genes are non-allelic.

4. Conclusions

Genetics of resistance studied of F2 population using

Allelic Relationship between Lr9 and the Leaf Rust Resistance Gene in Kharchia Local Mutant of Wheat

690

Table 1. F2 segregation for reaction to leaf rust pathotype 77-5 in three different crosses.

Observed number of plants

Sr.

No. Crosses Total no.

of plantsResistant (O-2+)3

+ 4– 4Total Expected ratio X2 (Cal.) Probability (P)

1. (WL711 + Lr KLM4-3B) × WL711) 122 86 14418363:1 1.32NS 0.10 - 0.25

2. (WL711 + Lr9 × WL711) 126 89 16219373:1 1.28NS 0.10 - 0.25

3. (WL711 – Lr KLM4-3B) × (WL711 + Lr9) 101 91 5231015:1 2.29NS 0.05 - 0.10

Table 2. Adult stage disease reaction of F2 population.

Seedling reaction (WL 711 + LrKLM4-3B) × WL711 (WL711 + Lr9) × WL711 (WL711 – Lr KLM4-3B) × (WL711+ Lr9)

Adult reaction Number of plants Adult reactionNumber of plantsAdult reaction Number of plants

0, 0;, ;, 0 75 0 79 0 86

1, 1+, 2, 2+ TS 5 TS 4 TS 1

(R) 5S 9 5S 6 5S 4

10S 4 10S 4 10S 5

3+, 4 40S 4 40S 7 40S 3

(S) 60S 26 60S 23 60S 2

80S 2 80S 3 80S -

crosses (WL711 +Lr9) × WL711 and (WL711 + LrKLM4-

3B) × WL711 segregated in ratio of 3:1 for reaction to

pathotype 77-5 of leaf rust. This suggested that rust re-

sistance in these stocks is under the control, of single do-

minant genes. Further, to study allelic relationship be-

tween Lr9 and LrKLM4-3B, F2 population of the cross

(WL711 +LrKLM4-3B) × (WL711 +Lr9) was studied. A

segregation ratio of 15:1 demonstrate that the two genes

Lr9 and LrKLM4-3B are two different non-allelic genes.

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