Establishment And Analysis Of SSCP Markers For Cotton Important Agronomic Traits

Molecular markers such as restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD), amplified fragment length polymorphi sm (AFLP), and simple sequence repeat (SSR) have been developed and used in cotton (Gossypium spp.) for constructing linkage maps, gene mapping, and genetic diversity detection.However,these markers has low level of polymorphism on a basis within cotton genomewide,especially within cultivated upland cotton,therefore, The lack of genetic diversity within cultivated upland cotton (Gossypium hirsutum L.) has severely hindered the construction of genomewide linkage maps and their applications in genetics and breeding. Single strand conformation polymorphism(SSCP)is a new detection technique for molecular markers with high polymorphism,density and strong genetic stability that saturate the cotton genetic map. Therefore, Using SSCP analysis is very important for construction of the cotton genetic map, marker assisted selection(MAS) and so on.In our study,Using SSCP analysis,we develop cotton molecular markers on the basis of cotton differentially expressed genes related to fiber development and oil synthesis- related genes.Then SSCP analysis was performed in 17 BILs with polymorphic primer pairs. Molecular markers associated with phenotypic traits that were screened out through the association analysis is useful for cotton QTL Localization, positional cloning and MAS and so on.1. A total of 162 primer pairs were designed for genes that were differentially expressed among interspecific introgression cotton lines for yield and fiber quality traits. A total of 146 primer pairs amplified a clear, strong and single band each from the genomic DNA when assayed by 1.5% agarose gels. Using the SSCP analysis, 54 primer pairs(37.0%) yielded 116 polymorphic loci among the four G. hirsutum genotypes, while 45 pairs(30.8%) yielded 111 polymorphic loci among the four G. barbadense genotypes. The SSCP polymorphism was much higher between the two species where a total of 79 pairs(54.1%) produced 260 polymorphic markers. Meanwhile,36 pairs(24.7%) yielded polymorphism between four G. barbadense genotypes and four G. hirsutum genotypes, respectively. The polymorphic SSCP markers grouped the eight genotypes into two species groups(G. barbadense genotypes and G. hirsutum genotypes), as expected.2. All ESTs of three oil synthesis- related genes that encode lysophosphatidic acid acyltransferase(LPAAT),diacylglycerol acyltransferase(DGAT) and phosphoenolpyruvate carboxylase (PEPC)were obtained from NCBI et al.A total of 75 primer pairs were designed for contigs that were consist of these ESTs by CAP3,respectively.A total of 55 primer pairs amplified a clear, strong and single band each from the genomic DNA when assayed by 1.5% agarose gels. Using the SSCP analysis, 21 primer pairs(38.2%) yielded 47 polymorphic loci among the four G. hirsutum genotypes, while 16 pairs(29.0%) yielded 30 polymorphic loci among the four G. barbadense genotypes. The SSCP polymorphism was much higher between the two species where a total of 39 pairs(70.9%) produced 104 polymorphic markers. Meanwhile,13 pairs (23.6%)yielded polymorphism between four G. barbadense genotypes and four G. hirsutum genotypes, respectively. The polymorphic SSCP markers grouped the eight genotypes into two species groups(G. barbadense genotypes and G. hirsutum genotypes), as expected.3. The association analysis between SSCP markers and 10 agronomic traits by SAS 6.0 showed that 63 markers associated with important agronomic traits of cotton were screened out include 24 markers(P<0.01) and 45 markers(P<0.05).Meanwhile, a total of 22 markers were associated with more than 2 agronomic traits. SSCP markers associated with important agronomic traits by association analysis provide a new foundation for map-based cloning and MAS.