Cloning And Bioinformatics Analysis Of Flanking Sequences Of Involved In Taxol Side-chain Biosynthesis

Taxol, extracted from taxus, is one of the most important natural anti-cancer drugs. Bacause of its unique anti-cancer mechanism, taxol has become the representative of a new generation of anti-cancer drugs. But chemical synthesis of taxol is inefficient and the extraction from plant will be a waste of resources. So biosynthesis of taxol has become the most promising method, but a few biosynthesis–ralated genes have not heen cloned. In this paper, we analyzed the chromosome distribuion of three genes (PAM,BAPT,DBTNBT) which involved in taxol side-chain biosynthesis.We have cloned three genes and their flanking sequences by homology cloning and thermal asymmetric interlaced PCR. Now we have obtained PAM and its flanking sequence 4.5kb, BAPT 6.6kb, DBTNBT 4.3kb.Analysis of the sequences by bioinformatics has been carried out. The results showed that: the ORF,promoter and terminator of PAM, which contains two exons, inturn is 1815bp,353bp and 723bp; the ORF,promoter and terminator of BAPT, which contains two exons, inturn is 1333bp,103bp and 245bp; the ORF,promoter and terminator of DBTNBT, which contains two exons, inturn is 1326bp,455bp and 158bp.46 taxol genes'base composition is analyzed and their G/C content is 43.4%. Codon preference analysis showed that the codons for Ala, Asp, Phe, Gly, His, Asn, Arg, Thr, Tyr have distinctly different frequency, which all prefer to use codons containing more A/T. Simultaneously the high-frequency codons also contain more A/T, which are identical with taxol genes'G/C content.The research of the three genes and the codon preference in this paper will lay a foundation of further study on the biosynthesis of Taxol side-chain.