The Cloning Of Genes Related To Cytoplasmic Male Sterility In Petaloid Type Carrot By CDNA-AFLP And CAPS

CMS (Cytoplasmic Male Sterility) is the base of crop heterosis. It is excellent material on studies of cytoplasm inheritance, G-CMS and pollen growth that has become the one of prevalence in field of molecular biology and gene engineering today. Lots of experimentations proved that they depend on each other for existence and their reciprocity take on important effect on development though cytoplasm is nucleus–independent. At present sufficient utilization of CMS has been hindered in carrot breeding due to be lacking in in-depth study and understand about form and mechanism of CMS in carrot and that utilizing of carrot heterosis has still been laggard. Therefore, it is indispensable to clone more gene related to CMS in carrot so that the basal theory would be provided to research mechanism of G-CMS. The new materials of sterility in carrot that created with gene engineering would be applied to production ultimately.cDNA-AFLP and BSA were dveloped to study differential expression in course of flower development on male sterile line and fertile line in CMS carrot . The genes related to CMS were cloned and analyzed comparably. In addition to, the atp6 genes of male sterile line and fertile line were obtained by CAPS marker respectively and their difference together with effect on CMS in carrot were analyzed. The main results were as follows:1. Three obvious cDNA differential fragments related to CMS were obtained successfully from selective PCR amplification with 128 pairs E+2/M+3 primer combinations by screening out differences of sterility and fertility together with phases.2. Sequence analysis showed that cDNA fragment of BY1949 is 364bp that coding 120 amino acid residues. It has 87% identity with proton-dependent peptide transporter-like protein and membrane protein from Arabidopsis thaliana as well as 40% identity with Nef1 from Oryza sativa. The retrieval of available literature indicated that its coding product affects lipid accumulation in the plastids of the tapetum and exine formation of pollen that resulting in male sterility in Arabidopsis thaliana.3. cDNA fragment of BY2562 is 432bp that coding 88 amino acid residues. It is over 90% identity with mitochondrial function protein of many vegetable consist of CMS carrot, CMS common sunflower, rape and Arabidopsis thaliana etc. And it has mitochondrial membrane protein YMF19 motif, which be involved in cytoplasmic male sterility (CMS) in Brassica by GenBank database.