Construction And Application Of Molecular Detection Technology System In Cotton Based On SSR Molecular Markers

Elite cotton varieties play a significant role on the quality and yield.The identity and purity of cotton variety have an significant effects on the quality and yield,which are important evaluation index for seed quality.With the number of varieties increasing,the method for identification field phenotypic agronomic traits has been difficult to meet the needs of the varieties identification,but,the molecular marker detection technology can meet the needs of the cultivars identification.This study based on SSR markers,combining the practical supervision of seedand theoretical research together.Through innovating rapid DNA extraction method,confirming the core SSR primer,ensuring the technology platform for electrophoresis detect,and confirming the cotton identification on purity and identity,in order to invent simple,low cost,accurate and high efficient molecular detection technology system which is of great significance to ensure the quality of cotton seeds.The main results were as follows:1.Through the screening of SSR primer polymorphism to ensure that 26 pair of core SSR primers were concluded,which covered the whole cotton chromosomes.The number of band patterns produced by each primer pair ranged from 3 to 11,with an average of 5.62.2.Through comparing the two silver stain methods of the non-denatured PAGE or the denatured PAGE,we find that denatured PAGE silver stain test can detect the real allelic variation,with clear stripe and the reliable result,which are suitable for detecting varieties identity,genetic relationship identification,etc.The non-denatured PAGE silver stain test is relatively simple,low cost and high detection flux,despite the presence of nonspecific banding,but does not affect detecting the molecular marker genotype,which are suitable for detecting varieties purity and molecular genetic diversity research,etc.3.Through the study of cotton purity with core SSR primers,we found three kinds of non pure type in molecular level.?:The cotton varieties with more non pure SSR loci and the loci are concentrated on only some special individuals.This type of varieties should belong to high generation with a small amount of mixed breed in the production.Ⅱ:The cotton varieties with less non pure SSR loci and the loci distribute randomly.This type varieties should be lack of breeding generation in the production.Ⅲ:The cotton varieties with more non pure SSR loci and the loci are derived from different genotypes.This type of varieties should belong to low generation material in the production,which have poor SSR loci and the plant phenotypic traits was unstable.The study ensures that the main factors for influencing the molecular purity are the non homozygous SSR markers loci rate and the non homozygous SSR marker loci homozygous rate.Combined with 26 of core SSR primers,this study proposed a variety of molecular purity calculation method:(varieties molecular purity =(1-1/26 [∑(SSR markers loci different genotype frequency)])* 100%).On the basis of crop seed quality standard,the study determine cotton molecular quality evaluation standard with 95% purity.4.This study research on authenticity determination methods combined the result of phenotypic traits in the field.Combined the 26 pair of core SSR primers,the study ensured the method to detect typical plant individuals,the fingerprint construction using typical plant individuals.Determined the identity of cotton varieties decision rules:(1)if the similarity coefficient of tested samples and contrast samples is 1,the tested samples and contrast samples can consider as the same species.(2)if the tested samples and contrast samples similarity coefficient between 1 and 0.98,the tested samples and contrast samples can consider as similar varieties.(3)if the similarity coefficient of tested samples and contrast samples < 0.98,the tested samples and contrast samples can consider as different species.5.This study manage cotton variety regional test successfully in shandong province using molecular detection technology,and research lumianyan 29 or 34 cotton varieties seed quality management.The study also provided an effective detection means for shandong province seed station,which also established a effective identification technology system for detecting lumianyan 29 and 34 cotton varieties.