| Pepper is a worldwide vegetable with great important to flavoring industry. It is the second large-area planting vegetable crop in China. EST (Expressed sequence tag) is the5’or3’ends of partial cDNA obtained from the sequencing of the clone which is selected from the cDNA bank and reflect the diversity of related gene in the population.With the rapid development of sequencing technology and bioinformatics, pepper ESTs are increased quickly in NCBI. Therefore, it is important that pepper ESTs are large-scale analysed via bioinformatics methods. But it is relatively weak that the application of EST database in pepper. In this research,116,952pepper ESTs were analysed with bioinformatics methods and EST-SSR makers were developed and the sequences function was analysed with Blast2Go. Furthermore, genetic diversity of17pepper cultivars and CMV resistance in pepper were analysed using EST-SSR. These studies will put a firm foundation for constructing high density genetic mapping, tagging genes, comparing genomics analysis, gene network analysis, and molecular evolution.The details results as following:1. Characterization and function annotation of EST-SSR in pepperDeveloped with a total of116,952pepper EST sequences from NCBI database, there were1,658SSR-containing EST identified by using SSRLoctor, the frequency of EST-SSR was1.92%. The most abundant motif types was dinucleotide motif (56.4%) and the followed by tri-(23.2%), tetra-(5.12%), hexa-(5.53%) and penta-nucleotide (9.71%). The top six motif types with high frequency included GA/TC (23.3%), AG/CT (17.0%), AT/TA (7.49%), TA/AT (5.01%), AGA/TCT (2.48%) and AAT/ATT (2.25%). Based on the1658SSR-EST, a total of810primer pairs were successfully designed and then annotated. The annotation results show that the number of cellular component, biological process and molecular function sequence was297,271and316. The annotation results divide into30types in the level2and one EST may annotate many functions. 2. EST-SSR analysis of genetic diversity in pepperGenetic diversity of17pepper cultivars were analysed by using100primers pairs. The results showed that80primer pairs were amplified effectively and46markers exhibited polymorphism with1～3alleles, and average of1.9alleles. These17pepper cultivars were divided into six groups by NTSYS2.10based on the similarity coefficient at0.718, the genetic relationship of17cultivars were related to their origin. It was revealed that the genetic relationship of accessions tested. The results indicated that the developed SSRs from ESTs of Capsicum were available and provided valuable evidence for the classification, conservation and utilization of the pepper germplasm.3. EST-SSR analysis of CMV resistance in pepperA resistant group and a sensitive group were divided with "BSA" method to search for the markers with resistance linked to CMV.80primers were used to select resistant markers among VC16a, SS69, the resistant group and the sensitive groups. After performing in EST-SSR analysis,65.0%EST-SSR primers were amplified effectively with1～5alleles, only one EST-SSR marker Y6158was identified to be linked to the resistant gene and was determined to be22.4cM from the resistant gene. Its application would accelerate our further research in molecular mechanism of pepper cucumber mosaic virus. |
PDF Full Text Request