Management of disease and insect resistance- methodologies and approaches
1. MANAGEMENT OF DISEASE AND
INSECT RESISTANCE
METHODOLOGIES AND APPROACHES
SUBMITTED BY:
Chavan Sonal
RAD/2020-24
Dept. of Genetics and Plant Breeding
Course Title: Breeding for Biotic and Abiotic Stress Resistance (GP-510)
SUBMITTED TO:
Dr. C.V. Sameer Kumar
Professor
Dept. of Genetics and Plant Breeding
2. RESISTANT VARIETIES
• Resistance is the ability of a plant variety to restrict the growth and/or development
of a specified pest and/or the damage it causes when compared to susceptible plant
varieties under similar environmental conditions and pest pressure.
• The cultivation of resistant varieties has been recognized as the most effective, ideal
and economical method of reducing crop losses (Stakman and Harrar 1957).
Collection of natural variability followed by finding
out the sources of resistance.
Incorporate the resistance gene (s) from
the donor parent(s) using various
methods.
Resistance breeding programmes:
In view of the dynamic nature of parasites, the resistant gene(s) fall susceptible after a few years
or are no longer effective. Therefore, the resistance breeding programme is a continuous one.
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3. Management of Disease and Insect Resistance
• The mode of inheritance of resistance in the host may be race-specific (vertical) or
nonspecific (horizontal).
• Depending on the mode of inheritance, several methods have been proposed for
the better utilization of the resistance gene(s).
• These techniques can prolong the average life span of resistant gene(s) and at the
same time reduce the risk of catastrophic losses.
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4. Feature Vertical Resistance Horizontal resistance
Pathotype specificity Race specific Race nonspecific
Nature of gene action Oligogenic Polygenic; rarely oligogenic
Response to pathogen Usually hypersensitive Resistant response
Phenotypic expression Qualitative Quantitative
Stage of expression Seedling to maturity Expression increases as
plant matures
Selection and evaluation Relatively easy Relatively difficult
Risk of ‘boom and burst’ Present (rarely durable) Absent (durable)
Suitable for a. Host b.
Pathogen
Annuals but not perennials.
Immobile pathogens, but for
mobile air-borne pathogens
Both annuals and
perennials.
All pathogens
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5. Feature Vertical Resistance Horizontal Resistance
Need for specific
deployment of resistant
varieties
Critical for success with
mobile pathogens
None
Need for other control
measures
Likely Much less likely
Host-pathogen
interaction
Present Absent
Efficiency Highly efficient against
specific races
Variable, but operates
against all races
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6. Horizontal Resistance
• Breeders and pathologists have preferred to utilize the specific (VR) resistance in the past,
because of the ease with which it can be incorporated into varieties, due to its simple
inheritance (often a single gene) and easily recognizable character.
• To produce more stable resistance, breeders rely on the incorporation of several major genes
into a plant variety. A variety with incorporated multiple major genes for resistance will not
become susceptible to the pathogen as quickly as one with a single major gene, because the
probability that several mutations or recombinations occur simultaneously is rather remote.
In practice, the incorporation of several major genes is only useful if these genes are at
different loci.
• Horizontal resistance (HR) reduces the apparent infection rate.
• HR provides a fairly stable resistance in preventing epidemics of plant diseases. Breeding for
durable horizontal resistance has been carried out successfully with potatoes resistant to late
blight and for diseases such as coffee berry disease caused by Glomerella cingulata, coffee
leaf rust caused by Hemileia vastatrix, wheat diseases caused by Helminthosporium
sativum, Fusarium, Colletotrichum graminicola, Septoria nodorum, powdery mildew,
and barley yellow dwarf virus. Other plants in which horizontal resistance has been
developed include chickpeas and broad beans.
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7. Management of Resistance Gene(s)
Recycling and Sequential Release of Resistance Gene(s)
Pyramiding of Resistance Gene(s)
Regional Deployment of Resistance Genes
Chromosome or Genome Substitutions
Multi-Line Cultivars
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8. Recycling and Sequential Release of Resistance Gene(s)
• Based on the same principle of crop rotations to control certain soil-borne or root-
infecting pathogens.
• Stevens (1949) suggested that a system of variety rotations, should be followed so that
inoculum of a particular race(s) does not build up in sufficient quantities over a period
of time to create an epidemic.
• The varieties rotated should, however, have different genes for resistance.
• After 5 or 10 years of
widespread planting of a new
host variety, it may be that the
formerly well-known races or
certain diseases will have become
scarce, and that the older host
varieties can be replanted with
profit.
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9. Proposal for race prediction and gene
rotation to keep development of new
disease resistant varieties for farmers
ahead of the development of new races
of disease organism (adapted from IRRI,
1980).
P = specific gene for pathogenicity in the
pathogen population;
p = allele of P incapable of being
pathogenic;
R = specific gene for resistance in a rice
variety;
r = allele of R resulting in susceptibility;
P = broken circle designates presumed
fate of P gene in the pathogen
population as disappearing or becoming
much reduced in frequency
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10. Steps involved:
Identify the monogene for resistance that is effective against races of the pathogen present in a
specific cropping area.
Use of monogene for resistance to develop varieties for the specific cropping area and release
these varieties. This will reduce and possibly eliminate the presently occurring races in the cropping
area.
Test the resistant monogene at sites remote from the specific cropping area, through international
testing programme. The race of pathogen that overcomes the monogene in one or more of the
remote areas will be representive of the race that will eventually evolve in the cropping area
following the introduction of the new varieties containing the monogene.
Identify, based upon results obtained from the programme (step 3), a monogene for resistance to
the race predicted to occur in the cropping area and use it in the variety development programme.
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11. • Rao (1968) noted that the system proposed by Stevens may be worth
consideration only when:
1. The old varieties are resistant to new races and new diseases that have become
important
2. The populations of the old races are not high at the time of replanting old
varieties
3. The new varieties are not superior to the old varieties in yield, quality and other
agronomic attributes; and
4. Seed multiplication and distribution is not cumbersome.
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Since in an on-going breeding programme improvements in cultivars are
always being made, the utility of the system proposed by Stevens may be
limited.
However, if a judicious back-cross programme is followed keeping the
original genotype, with additional genes for resistance added, as new races
came up, the system could be balanced.
Release one gene for resistance and wait until it becomes ineffective;
release the second gene and so on.
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12. Examples:
Control stem rust of wheat in Australia between 1938 and 1950 (LA. Watson and Luig).
Resistance to brown plant hopper (Nilaparvata lugens) at IRRI.
• IR 26 and IR 1561-228-3 varieties of rice with resistant gene Bph 1 were released in 1973 and
1974 respectively.
• Towards the end of 1975 and in 1976, these varieties started to show susceptibility at some
locations in the Philippines.
• But by that time multiple disease- and insect-resistant varieties with bph2 resistance gene for
BPH had become available (IR36 and IR38) and were released as replacements for the
varieties with Bph 1. Later on at IRRI, breeding lines with Bph3 and bph4 for resistance to
BPH were available. A proposal for race prediction and gene rotation to keep development
of new disease-resistant varieties of rice for farmers ahead of the development of new races
of disease organisms was advocated at IRRI (1980).
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13. Pyramiding of Resistance Gene(s)
• Simultaneous introduction of diverse genes for resistance into the cultivar was
proposed for the first time by I.W. Watson and Singh (1952).
• Athwal (1953) gave a scheme for introducing two genes, following a system of back-
crossing and taking advantage of epistatic reaction types.
• The system is based on the mutation in the pathogen at more than one locus being
much rarer than the mutation at one locus. The higher the number of diverse genes
introduced into a single variety, the greater would be the longevity of its resistance.
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14. • A pyramid could be constructed with major genes, minor genes, effective genes,
ineffective genes, race-specific genes, non race-specific genes, or any other type of
host gene that confers resistance.
CONVENTIONAL TECHNIQUE:
Serial gene pyramiding: Genes are deployed in same plant one after other.
Pedigree breeding
Backcross breeding
Recurrent selection
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18. Advantages of Gene pyramiding:
1. Widely used for combining multiple disease resistance genes for specific races of a
pathogen.
• Pyramiding is extremely difficult to achieve using conventional methods - Consider
- phenotyping a single plant for multiple forms of seedling resistance – almost
impossible.
2. Important to develop 'durable' disease resistance against different races.
3. Mainly used to improve existing elite cultivar.
4. Eliminates extensive phenotyping.
5. Control linkage drag.
6. Reduces breeding duration.
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19. CRITICISM:
If a race which can attack the combined resistance is produced, then different resistant
genes became susceptible simultaneously.
Flor (1958) reported simultaneously induced mutations for virulence involving two loci in
the fungus Melampsora lini. Of course, we do not know how much occurs in nature and
whether more loci can simultaneously be affected.
Schafer et al. (1963) pointed out the difficulty in this system of detecting the individual
genes for resistance in a single genetic stock, because of their common protection against
available races of the pathogen.
Main factors affecting gene pyramiding
1. Characteristics of the target traits/genes.
2. Reproductive characteristics.
3. A breeder's capability to identify the 'desired' genotypes.
4. Operating capital.
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22. Regional Deployment of Resistance Genes.
• Resistant varieties with different resistance genes should be developed and
recommended for different geographical regions of the country, where the crop
covers a sizable area. As pointed out by R.R. Nelson (1972), this type of gene
deployment is essentially a geographical multiline. A formal plan for regional
deployment of genes is in effect for resistance.
• When a number of genes are in operation, the possibility
of build-up of a super-race is minimized, and the pathogen
will became stabilized. A similar line of work with Sr and Yr
isogenic lines will help in the release of stable varieties for
all the three prevalent wheat rusts in India.
•This approach can be followed for any other crop
diseases and insects when enough genes are identified.
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23. • M.S.s. Reddy and M.V. Rao (1979) suggested a strategy for controlling leaf rust, Puccinia
recondita f.sp. tritici, of wheat in India by the use of regional deployment of resistance genes.
India could be divided into three regions, (A) central plains, (B) northern Himalayan, and (C)
southern Nilgiri and Palani Hills. Stronger genes (Lr 9 or Lr 19 or Lr 10 + Lr 15) can be deployed
in region A, Lr 1 + Lr 10 + Lr 1 b or Lr 3 ka + Lr 10 + Lr 17 in region B, and Lr 3 ka + Lr 10 + Lr 20
or Lr 1 + Lr 10 + Lr 17 in region C.
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24. Chromosome or Genome Substitutions
• For breeding resistant varieties, the sources of disease resistance are sought and the
resistant gene(s) are incorporated in cultivated varieties. If gene(s) for resistance are
not available in the cultivated species, the breeder transfers resistance from related
species/genera.
• Due to genetic or cytoplasmic or genome cytoplasm interactions, the chromosomes of
different species fail to pair either completely or partially, resulting in sterility of Fl. To
facilitate recombination, either the F 1 or the amphidiploid is back-crossed with the
recurrent parent. In this way the whole genome or a whole chromosome or a
segment of a chromosome of a recurrent parent is replaced by the donor parent.
• The genome of a wild or related species (donor parent) could be incorporated into
the susceptible cultivar of a cultivated species (recipient) through hybridization and
chromosome doubling, using colchicine. By using colchicine, amphidiploids are
produced which have the full normal diploid genome of both the species.
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25. Resistance to clubroot
(Plasmodiophora brassicae) has
been transferred from the turnip
(Brassica campestris) (2n = 20,
AA) to the Swede turnip (B. napus)
(2n =36, AACC).
Johnston (1974) used colchicine
to double the Fl hybrid to produce
an amphidiploid which can be
maintained indefmitely and is
more easily back-crossed to B.
napus. Resistant second back-
cross progenies were virtually
identical with the recurrent
parent.
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26. • Ideally, the alien chromosome
segment should be as small as
possible, particularly if the two species
show extensive chromosomal
differentiation. Otherwise, the
substitution of an alien chromosome
segment for a chromosome segment
of the recipient species may result in
undesirable duplications, deletions
and linkages of genes. The effect is
large in diploids, but is often reduced
in polyploids, where the effect of
single chromosomes is smaller
because of the duplication of genetic
material in two or more different
genomes.
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28. Multi-Line Cultivars
• Multiline varieties are mixtures of several purelines of similar height, flowering and
maturity dates, seed colour and agronomic characteristics, each of which has a
different gene for resistance to the given disease.
• The purelines constituting a multiline variety must be compatible i.e., they should
not reduce the yielding ability of each other, when grown in a mixture.
• The idea of multiline varieties was put forward by Jensen in 1952 for use in
cereals.
• A multiline cultivar is a population of plants that is agronomically uniform but
heterogeneous for genes that condition reaction to a disease organism.
• It has long been recognized that crop homogeneity is a condition favouring
epiphytotics. The use of multiline varieties in self-pollinating crops has been
advocated since the late nineteenth century.
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29. Multiline cultivars
Clean crop approach
• All component lines of the mixture
would be resistant to all prevalent
races of the disease to be controlled
• The aim of this scheme is to keep the
crop as free of disease as possible,
and at the same time to reduce the
threat of catastrophic disease losses
following shifts in the racial com
position of the pathogen population
Dirty crop approach
• Each line in the mixture also carries a
different single gene resistance;
however, none of the lines is resistant
to all known races of the pathogen
The programmes of multi-line production are based on two radically different
philosophies for disease control (Marshall 1977).
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30. Development of Multiline Varieties
• Two main steps in the development of multilines:
1. Development of Component lines
• Identification of several sources of distinct and preferably, known genes for
resistance to the concerned disease.
• These resistance genes are transferred into a elite variety or line to produce as
many near isogenic lines -done by conventional backcross (5-6 backcrosses), a
limited backcrossing (2-3 backcrosses, followed by pedigree selection).
2. Evaluation and Grouping of Component lines
• Evaluation- multilocation trails followed by evaluation for compatibilty or nicking
ability
• Grouping – the number of component lines should be large, 15-20 according to
Borlaug (1959), if durability of resistance is desired.
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31. The prerequisities of multiline cultivar approach are (M.V. Rao):
• Proper identification of diverse genetic sources of resistance.
• Adequate race survey.
• Desirable and commercially acceptable.
• A widely adapted recurrent parent.
• By following a conventional or limited back-crossing programme (the latter was
found to be more beneficial by Borlaug because of the transgressive segregants,
which were better than the recurrent parents), phenotypically similar but
genotypically different lines are developed by crossing the recurrent parent with
stocks carrying diverse genes for resistance.
• All these phenotypically similar lines are mixed together and distributed to the
farmers as a composite variety. If a line/genotype is affected by a new race, it is
withdrawn from the composite bulk.
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32. Mechanism of Action of Multi-Lines
Reduction of Xo and r (Vander Plank 1963).
• The pathogen increases from initial inoculum (Xo) at the rate r in time t, and
results in x proportion of susceptible tissue becoming infected.
• A variety with vertical resistance (VR), being selectively resistant to the race
population, reduces Xo but r remains unchanged and is usually high (not
limiting); x may be very large by the end of the disease season. Therefore, VR is
valuable only as long as it gives resistance to all prevalent races and keeps Xo
very small for all.
• A variety with horizontal resistance (HR), i.e. resistant to all races of the
pathogen, does not reduce Xo, but reduces r. Since r is small, the rate of
epiphytotic development is reduced to the point where the host matures with
small x and little measurable damage.
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33. Advantages of Multi-Lines
• They provide a mechanism of "synthesized horizontal resistance" which unlike pure line
cultivars can utilize several resistant genes.
• They extends the life of a given resistant gene(s) and enable a resistance breeding
programme to be reduced in size, while the breeder carries on parallel improvement in the
recurrent parent.
• The use of a multi-line cultivar would stabilize the yield to optimize production on a given
farm.
• This reduces the risk of homogenizing the pathogen population on a global scale.
• Multi-line cultivar may out-yield the recurrent parent even in the absence of rust infection.
• Multi-lines hold great promise as a dynamic and natural biological system in effectively
balancing the relationship between the host and the pathogen.
• Criticisms of the Multi-Line Approach: The multi-line approach of management of VR genes
for controlling pathogens with great epiphytotic potential such as rusts has been criticized as
expensive (Suneson 1960), agronomically conservative (R.M. Caldwell 1966 and Hooker
1967a), a breeding ground for new races (Hooker 1967, Simmonds 1962, Vander Plank 1963),
and possibly even a super-race (R.M. Caldwell 1966).
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34. Multiple Resistance
• Resistance to two or more diseases has been bred into individual cultivars since.
• One of the approaches for incorporating the multiple resistance could be the
screening for multiple resistance line(s) from germplasm followed by crossing and
screening for more than two diseases and/or insect pests in the segregating
generations.
• W.A. Orton (1909) succeeded in
combining resistance to Fusarium wilt
and root knot nematode in cowpea and
cotton. Hope and H44 wheat cultivars
combined resistance to leaf rust, stem
rust and covered smut. Multiple
resistance has long been a breeding
achievement in tobacco, sugarbeet, corn,
beans, and many other crops.
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35. Transgenics and Cisgenics
• Modification of transgenic
classifications, for example, the
concept of cisgenics (allowing the
addition of genes from a crossable
species) as opposed to transgenics
(the addition of a gene or genes
from a non-crossable species), may
increase the workable space in crop
modification.
• Success of many of the advances in
engineering disease resistance in
crop species, of course, depends on
societal acceptance of various
approaches to plant genome
modification.
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36. CRISPR–Cas9
• Strides in genome editing, particularly
the CRISPR–Cas9 system, have increased
interest towards the development of
disease resistance through the
modification of susceptible (S) genes of
the plant.
• The classic example is mlo, a recessive R gene
of barley with resistance toward powdery
mildew. The null allele provides broad,
durable resistance against the pathogen
Blumeria graminis f. sp. hordei. Simultaneous
editing of all three homoeoalleles of MLO
locus in hexaploid wheat conferred recessive
resistance against powdery mildew in one
generation.
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37. • Disruption of downy mildew resistance-6 (DMR6) was originally identified in
Arabidopsis and suppresses free salicylic acid (SA) levels. Enhanced SA levels are
associated with reduced susceptibility to a variety of pathogens, particularly
bacterial pathogens. Mutations created by CRISPR–Cas9 in DMR6 orthologs in
tomato were reported to confer resistance to a number of pathogens, including P.
syringae pv. tomato, Phytophthora capsica, X. perforans, and Xanthomonas
gardneri.
• The ease of multiplexing, i.e., the simultaneous targeting of several genes with a
single molecular construct, is one of the major advantages of CRISPR/Cas9
technology with respect to MN, ZFN, or TALEN.
• Example: the simultaneous mutation of 14 different genes by a single construct
has been demonstrated in Arabidopsis (Peterson et al., 2016). In crops, several
multiplex genome editing (MGE) strategies were reported early on (Ma et al.,
2014; Xing et al., 2014; Zhou et al., 2014; Xu et al., 2016), which were all based on
a common strategy, i.e., the assembly of multiple gRNAs under the control of a U3
or U6 promoter into a single construct.
• In maize, the ISU Maize CRISPR platform (Char et al., 2017) permits the cloning of
up to four gRNAs for multiplex gene targeting.
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38. Challenges
• The challenge is to demonstrate that the promises made by proofs of concept in
confined environments can be maintained under field conditions.
• Most of the genes inactivated by CRISPR/Cas9 technology in order to obtain
disease resistance are likely to have roles in the physiology of the plant other than
that linked to the life cycle of the pathogen.
• For example, triple knockouts of wheat TaMLO were not only resistant to
powdery mildew but also showed leaf chlorosis (Wang et al., 2014), whereas
EMS-induced triple mutants with non-conservative point mutations in TaMLO did
not show obvious pleiotropic phenotypes (Acevedo-Garcia et al., 2017). Therefore,
encouraging greenhouse observations of plant development or measurements of
key parameters such as height, leaf area or grain weight absolutely must be
confirmed under field conditions by multi-environmental yield trials in order to
measure the relative importance of negative side effects.
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40. • The varieties ASD 16 and ADT 43 are the two popular, high yielding, and widely grown rice cultivars of South India,
which are susceptible to bacterial blight (BB), blast, and sheath blight diseases.
• The present study was carried out to improve the cultivars (ASD 16 and ADT 43) through introgression of bacterial
blight (xa5, xa13, and Xa21), blast (Pi54), and sheath blight (qSBR7-1, qSBR11-1, and qSBR11-2) resistance genes/QTLs
by MABB (marker-assisted backcross breeding).
•IRBB60 (xa5, xa13, and Xa21) and Tetep (Pi54; qSBR7-1, qSBR11-1, and qSBR11-2) were used as donors to
introgress BB, blast, and sheath blight resistance into the recurrent parents (ASD 16 and ADT 43).
• Homozygous (BC3F3 generation), three-gene bacterial blight pyramided (xa5 C xa13 C Xa21) lines were developed,
and these lines were crossed with Tetep to combine blast (Pi54) and sheath blight (qSBR7-1, qSBR11-1, and qSBR11-
2) resistance.
• In BC3F3 generation, the improved pyramided lines carrying a total of seven genes/QTLs (xa5 C xa13 C Xa21 C Pi54 C
qSBR7-1 C qSBR11-1 C qSBR11-2) were selected through molecular and phenotypic assay, and these were evaluated
for resistance against bacterial blight, blast, and sheath blight pathogens under greenhouse conditions. We have
selected nine lines in ASD 16 background and 15 lines in ADT 43 background, exhibiting a high degree of resistance to
BB, blast, and sheath blight diseases and also possessing phenotypes of recurrent parents.
•The improved pyramided lines are expected to be used as improved varieties or used as a potential donor in
breeding programs. The present study successfully introgressed Pi54, and qSBR QTLs (qSBR7-1, qSBR11-1, and
qSBR11-2) from Tetep and major effective BB-resistant genes (xa5, xa13, and Xa21) from IRBB60 into the
commercial varieties for durable resistance to multiple diseases.
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41. Bacterial blight:
IRBB60 (xa5, xa13, and Xa21)
Blast
Tetep (Pi54)
Sheath blight
Tetep(qSBR7-1, qSBR11-1, and qSBR11-2)
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44. • Brown planthopper (BPH) is a major insect
pest of rice that causes 20–80% yield loss
through direct and indirect damage.
• The identification and use of BPH
resistance genes can efficiently manage BPH.
• A molecular marker-based genetic analysis
of BPH resistance was carried out using 101
BC1F5 mapping population derived from a
cross between a BPH-resistant indica variety
Khazar and an elite BPH-susceptible line
Huang–Huan–Zhan.
A total of 702 high quality polymorphic single nucleotide polymorphism (SNP) markers, genotypic data, and precisely
estimated BPH scores were used for molecular mapping, which resulted in the identification of the BPH38(t) locus on
the long arm of chromosome 1 between SNP markers 693,369 and id 10,112,165 of 496.2 kb in size with LOD of
20.53 and phenotypic variation explained of 35.91%. A total of 71 candidate genes were predicted in the detected
locus. Among these candidate genes, LOC_Os01g37260 was found to belong to the FBXL class of F-box protein
possessing the LRR domain, which is reported to be involved in biotic stress resistance. Furthermore, background
analysis and phenotypic selection resulted in the identification of introgression lines (ILs) possessing at least 90%
recurrent parent genome recovery and showing superior performance for several agro-morphological traits. The BPH
resistance locus and Ils identified in the present study will be useful in marker-assisted BPH resistance breeding
programs.
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45. To date, 37 BPH resistance genes have been identified from cultivated and wild species of
Oryza:BPH1, BPH2, BPH3, BPH4, BPH5, BPH6, BPH7, BPH8, BPH9, BPH10, BPH11(t), BPH12(t),BPH12,
BPH13(t), BPH14, BPH15, BPH16(t), BPH17, BPH18, BPH19(t), BPH20, BPH21, BPH22(t),
BPH23(t),BPH24(t), BPH25(t), BPH26(t), BPH27, BPH28, BHP29,BPH30, BPH31, BPH32,BPH33, BPH3
and BPH35, BPH36 and BPH37.
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