Development Of DNA Fingerprinting For The Hybridity Determination Of 'Shuza No.9' (Brassica Napus)

The main objectives of this study are to develop RAPD molecular markers in 'Shuza No.9' (Brassica napus), then convert the markers into SCAR molecular markers for evaluating hybridity of Fi hybrid.Both maternal-specific and paternal-specific DNA bands were amplified with the DNAs of parents of hybrid rapeseed 'Shuza No.9' by primer S18 (CCACAGCAGT) and S31 (CAATCGCCGT) which were selected from 50 10-mer RAPD primers analyzed in this study. Then the primers were used to amplify all the DNAs of F1 rapeseed by PCR , and the results indicated that the specificity and stability of primer S18 and S31 was trusty. The hybrid seed purity of 'Shuza No.9' could be determined reliably with them.In order to improve the reproducibility and reliability of PCR assays, the RAPD markers need to be en verted into SCAR markers. The two RAPD marker fragments, named S18750 and S31550, were cloned and the identity of these clones with respect to the corresponding RAPD markers were examined by bacterium PCR and plasmid PCR. 4 clones of S18750 and S31550 fragments come from parents and F1 hybrids amplified by RAPD PCR, were sequenced.According to sequencing results, two sequences of S18750 which one was cloned from paternal-specific DNA band, another was cloned from F1 specific DNA band are same sequence. Two sequences of S31550 which one was cloned from maternal-specific DNA band, another was cloned from F1 specific DNA band are same sequence too.Based result of sequence of S18750 fragment, designed a pair of primers namedS18750-1 and S18750-2 for male parents specific band, and same way for female with another pair of primers named S31550-1 and S31550-2. SCAR markers were developed. Using these SCAR primers to amplify random single F1 hybrid DNA, the parent specific bands could be amplified apparently. These results proved it was success that RAPD markers be converted to SCAR markers. The SCAR markers will be used to authentication of hybrid purity of 'Shuza No.9' rapeseeds from yields.Base of the DNA markers and a method using to extract DNAs from plants, a system in which hybrid purity of 'Shuza No.9' rapeseeds could be identified quickly, well and truly was build.