Cloning And Sequence Analysis Of Fragments Associated With Bolting In Chinese Cabbage

Chinese cabbage(Brassica rapa L.ssp.pekinensis ) is an important vegetable of Brassica in China.Early bolting is a major problem in Brassica rapa crops production.In spring,the temperature from low to high and the day time increase is the major reason of premature bolting in Chinese cabbage.It causes a great of loss in product.The effective solution is to breed late bolting variety.However,traditional breeding techniques is proved to be a laborious process with a certain degree of blindness,severely restricted the development of Chinese cabbage breeding technology and making late bolting breeding difficult in China. Therefore,exploring genes involved in vernalization response is important to Chinese cabbage varieties improvement.Here,one extremely early bolting line(DH-54) and one extremely late bolting line(DH-43) were employed,and the cDNA-AFLP approach was used to identify key components involved in the low-temperature required vernalization response. The main conclusions are as follows:1.Of 256 primer recombinations screened,a total of 191 differential expressed transcript-derived fragments(TDFs) were identified,and 82 TDFs were sequenced.BLAST and alignments showed that 52 candidate TDFs shared high levels of similarity with genes of known function,22 TDFs of unknown function and 8 novel ESTs.2.52 function knowed TDFs were classified using MIPS,The TDFs of known function were involved in metabolism,energy,cellular transport and transport facilitation and transport routes,cell rescue,defence and virulence,subcellular localization,expression regulator,signal transduction,Protein fate,cell cycle and DNA procession,regulation of metabolism and protein function,cell fate.3.Primer combinations for 14 randomly selected candidate genes were designed to do real-time PCR to analyze the gene expression pattern.The results showed that among these genes,six were up-regulated,three showed up-down-up-regulated,and five showed differential expression in two materials.In addition,considerable agreement was gotten from results of cDNA-AFLP and real time quantitative PCR.