| In view of the fact that Cruciferae crops not only account for the largest growing area of vegetable crops but also play an important role in agricultural activities and lives of people in China, it has been our objective that their germplasm resources were fully utilized. However, there have been many of ambiguities and divergences for the taxa of family, genus and species in Cruciferae for a long time. With the developments of molecular biology and bioinformatics, it is possible to elucidate the phylogenetic evolutionary relationship of Cruciferae at the molecular level. It was well known that studies on evolutions and taxa of species have been more and more fashionable by means of either some encoding and non-encoding genes of cpDNA and nrDNA or repeat sequences, ITS, and multiple genes families of nrDNA. Cytochrome P450 play a critical role in plant secondary metabolites synthesis as an ancient gene superfamily. In this paper, phylogenetic relationship of 13 species involved in 6 genera of Cruciferae were carried out through both the clones of homologous sequences with the primers designed on the basis of conserved regions of CYP86MF gene in cytochrome P450 gene superfamily and the differential analyses of them. Meanwhile, complete sequences of some genes in cytochrome P450 gene superfamily were isolated and identified by SMART PCR-RACE strategy, and expressed in E. colt. The results were as follows:(1) Isolated by PCR from 11 species of Cruciferae, eleven homologous gene segments that deduced amino acids were identities of over 80% at nucleotide sequence level and similarities of over 70% at amino acid sequence level.(2) The cytochrome P450 gene named BCCYP86MFS and isolated from Brassica campestris ssp. pekinensis cv. Huangyal4 was complete nucleotide sequence of 1 854 bp with a nucleotide ORF (1575 bp), which encoded a protein consisting of 524 aa with molecular weight of 62.2 kDa and pI of 8.96. Strongly basic (+) amino acids, strongly acidic (-) amino acids, hydrophobic amino acids and polar amino acids of the protein were 13.74%, 11.64%, 36.45% and 22.70% respectively, and predicted secondary structure of the protein revealed many conserved domains such as N-glycosylation site, protein kinase C phosphorylation site, casein kinase II phosphorylation site, N-myristoylation site, cAMP- and cGMP-dependent protein kinase phosphorylation site, tyrosine kinase phosphorylation site and a cytochrome P450 cysteine heme-iron ligand signature which was typical of cytochrome P450. a -helix and B -sheet of the protein is 47.7%, 45.0% respectively. Compared with cDNA, the BCCYP86MF5 genomic DNA being similarities of 95.2%and 85.9% to BCCYP86MFI and CYP86C4 at amino acid sequence level had no intron. The result of the Northern hybridization showed that it was expressed only in floral buds.(3) A full-length cDNA sequence of RSCYP86MF gene as one of cytochrome P450 gene superfamily from Raphanus sativus cv. Yuanbai was composed of 1 818 bp with a nucleotide ORF (1 581 bp), which encoded a protein of 526 aa with molecular weight of 62.2 kDa and pI of 8.96. Strongly basic (+) amino acids, strongly acidic (-) amino acids, hydrophobic amino acids and polar amino acids of the protein were 13.12%, 11.59%, 36.69% and 23.19% respectively. The secondary structure of this protein speculated had a typically conserved domain of cytochrome P450 cysteine heme-iron ligand signature (FNAGPRLCIG) and a hydrophobic domain at N-terminal of amino acid. In addition to the numerary variance, RSCYP86MF had the same domains as BCCYP86MF5 for the protein structure. Amino acid sequence of the protein with 47.7% of a -helix and 45.0% of B -sheet showed identities of 90%, 84% and over 38% to BCCYP86MF1, Atlgl3150 and every member of CYP86 family, respectively. The RSCYP86MF gene had no intron. The result of the Northern hybridization showed that it was expressed only in floral buds.(4) A complete cDNA sequence of OVCYP86MF gene as one of cytochrome P450 gene superfamily from Orychophrogmiis violaceus showed 1...
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