Gene pyramiding

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Gene pyramiding


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SEMINAR Gene Pyramiding OF QULITATIVE TRAITs Presented by Mr . AMBAVANE AJINKYA RAJENDRA M.Sc . (Ag) Regd no.-2153



Gene Pyramiding -Introduction:

Gene Pyramiding -Introduction Developing elite lines and varieties requires breeders to combine traits from multiple parents, a process called gene pyramiding or stacking. Pyramiding is the sequential accumulation of genes into a single line or cultivar. A pyramid could be constructed with major genes, minor genes, race-specific genes, non race-specific genes, or any other type of host gene that confers resistance. 4

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It includes Stacking of traits Stacking of events Stacking of genes A genetically modified organism (GMO) and all subsequent identical clones resulting from a transformation process are called collectively a transformation event. If more than one gene from another organism has been transferred, the created GMO has stacked genes (or stacked traits), and is called a gene stacked event . 5


OBJECTIVES OF GENE PYRAMIDING Widely used for combining multiple disease or pest resistance genes for specific races of a pathogen or insect. Important to develop durable resistance against different races. Main use- to improve existing elite cultivar. Reduces breeding duration. 6

Types of Gene Pyramiding:

Types of Gene Pyramiding Conventional technique Serial gene pyramiding : Genes are deployed in same plant one after other Molecular technique Simultaneous gene pyramiding : Genes are deployed at a time in a single plant 7

Gene-pyramiding scheme cumulating six target genes:

Gene-pyramiding scheme cumulating six target genes Hospital et al ., 2004 8

Fixation steps:

Fixation steps Production of doubled haploids from root genotype Crossing the root genotype with a blank parent and selfing the offspring Crossing the root genotype with one of the founding parent Selfing the root genotype 9

Different schemes of backcrossing for gene pyramiding.:

Different schemes of backcrossing for gene pyramiding. 10


A 11


INTRODUCTION Bacterial blight (BB) of rice caused by Xanthomonas oryzae pv oryzae ( Xoo ) Yield losses caused by bacterial blight in some cases can reach 50% A dominant gene Xa21 known to resistance to most races of BB ( Oryza longistaminata) Stem borer damage is a serious problem in rice, causing a 10–30% loss of total yield. Bacillus thuringiensis ( Bt ) produces a characteristic crystalline insecticidal protein Sheath blight disease is caused by the soil-borne fungal pathogen Rhizoctonia solani . Yield loss ranges from 8% to 50% Chitinase gene RC7 , isolated from an R. solani interrogated with rice cultivar showed enhanced resistance to sheath blight disease 12

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Result In the bioassay with infection of X. oryzae pv oryzae , different plants of the homozygous lines showed different levels of infection, but all of them showed a lesion length of less than 3.1 cm, which is considered to be resistant . When the plants of the homozygous lines were exposed to the infection of R. solani , observed a broad range in levels of infection. Average infection was approximately 40%, which represents 60% protection. In the bioassay of our final selected line with infestation with larvae of YSB, found that all the plants were completely resistant, showing 100% mortality of larvae. 14


B 15

The ancestry of SLG12 *Strain with single grain weight exceeding 50 mg.:

Taiho Cho- ko -to Aokei 79 Fusayoshi Nok - khao -Ngo F 1 813042* BG1* SLG1* SLG12 Hao Gang The ancestry of SLG12 *Strain with single grain weight exceeding 50 mg. 16

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Line SLG12 with an extraordinarily large size of grains was obtained from a series of crosses of lines selected for larger grain size BG1 (Big Grain 1) was selected from Taiho -Cho- ko -to as the first large-grain line which weighed more than 50 mg . SLG1 (Super large grain 1) with a single grain weight of more than 70 mg was developed from BG1/813042 SLG12 was obtained from SLG1/ Hao -Gang, the latter being a Chinese cultivar having a very large grain width. 17

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Variety 1 grain weigh(mg) Grain length (mm) Grain Width (mm) Koganebare 27.3 5.9 3.6 Nishihomare 28.4 6.3 3.5 BG1 56.4 9.7 4.3 SLG1 76.8 12.2 4.4 Hao Gang 47.9 8.6 4.6 SLG12 77.9 12.8 4.9 Grain size of SLG12 and some other varieties 18


C 19


INTRODUCTION Leaf rust is an important foliar disease of wheat. Two leaf rust resistance genes, Lr9 and Lr24 , have been pyramided through the use of simple sequence repeats (SSR) markers. The obtained wheat lines which carry the two resistance genes are indeed resistant to the leaf rust races . The lines have been used as parents to cumulate other resistance genes ( Lr22a ). This first cycle of ‘pyramidisation’ allowed us to evaluate the costs for marker-assisted selection (MAS). 20


METHEDOLOGY The Lr9 and the Lr24 resistance gene donors were backcrossed seven times to susceptible Swiss winter wheat cultivar Arina and selfed to produce the F8 generation. Two leaf rust resistance genes, Lr9 and Lr24 , have been pyramidised through the use of simple sequence repeats (SSR) markers After 6 years of classical breeding, MAS was applied to confirm the presence of Lr9 and/or Lr24 in 30 of the F8 remaining lines. 21

Leaf rust markers:

Leaf rust markers The MAS at F 2 with dominant PCR-based markers discard the plants without Lr genes Lines without the genes continue to appear after self-pollination of heterozygous plants. The second PCR was performed with accurate concentration of DNA isolated from 6 to 10 plants. For the heterozygous lines, the band intensity was lower than the one obtained with homozygous lines. The heterozygosity of these lines was confirmed by analyzing 6 plants separately. 30 lines tested in yield trials, 6 lines have markers for Lr9 and Lr24 , 10 lines only for Lr24 , 4 lines only for Lr9 and 10 lines have no markers for neither resistance genes. 22


RESULT The lines obtained, displaying good resistances and excellent bread making quality. But it had low yield and more difficulties to reach uniformity However, the lines have been used as parents to cumulate other resistance genes 23


ADVANTAGES OF GENE PYRIMIDING Development of resistance variety for new races of pathogen and insect Durable resistance It helps in crop improvement programme Reduce breeding duration 24


LIMITATION OF GENE PYRAMIDING Lots of efforts have to made to incorporate several major gene into single cultivar Pyramiding is extremely difficult to achieve using conventional methods. It Is very difficult to interrogate one gene from one cultivar or one species to another one by conventional methods . Consider - phenotyping a single plant for multiple forms of seedling resistance – almost impossible 25

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Stability of all desired gene in one plant is another issue which limited the “ Gene Pyramiding ” It requires extensive testing against various new races of pathogen Chances of formation of new race of pathogen 26


EXAMPLES OF GENE PYRAMIDING Pyramiding of Greenbug (Homoptera: Aphididae) Resistance Genes in Wheat Gene pyramiding for soybean mosaic virus resistance using microsatellite markers Gene pyramiding for powdery mildew resistance in wheat Pyramiding multiple genes for resistance to soybean mosaic virus in soybean using molecular markers 27

MAS based gene pyramiding for important traits in major crops.:

MAS based gene pyramiding for important traits in major crops. 28


Conclusion Gene pyramiding is an important strategy for germplasm improvement. Pyramiding requires that breeders consider the minimum population size that must be evaluated to have a reasonable chance of obtaining the desired genotype. Gene pyramiding with marker technology can integrate into existing plant breeding programmes all over the world to allow researchers to access, transfer and combine genes at a rate and with a precision. MAS based gene pyramiding has the potential To increase the rate of genetic gain when used in conjunction with traditional breeding 29


REFERENCES Balasubramanian P. (2003), Transgenic Bt Plants: Resistance Management Strategies. African Journal of Biotechnology 5 ( 10):781-785 Joshi R. K. and Nayak S. (2010), Gene pyramiding-A broad spectrum technique for developing durable stress resistance in crops . Biotechnology and Molecular Review . 5 (3):51-60. K. Datta , N. Baisakh , K. Maung Thet , J. Tu , S. K. Datta (2002) Pyramiding transgenes for multiple resistance in rice against bacterial blight, yellow stem borer and sheath blight. Theor Appl Genet 106 :1–8 Manyangarirwa W., Turnbull M., McCutcheon G. S., and Smith, J. P. ( 2006), Gene pyramiding as a Bt resistance management strategy African Journal of Biotechnology . 5 (10), pp. 781-785. 30

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Odile Moullet , Dario Fossati , Fabio Mascher1, Roberto Guadagnolo and Arnold Schori (2010). Use of marker- assisted selection (MAS) for Pyramiding leaf rust resistance genes ( Lr9 , Lr24 , Lr22a ) in wheat Tadashi Takita , and Norindo Takahashi (2003), A rice line with very largegrain obtained by pyramiding genic effects, http://www. rice_genetics /rgn5/v5 VI33.html 31

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