芜菁花青素合成酶基因的克隆、序列分析及表达

目的：克隆津田芜菁和赤丸芜菁花青素合成酶（ANS）基因并研究其表达特性。方法：用UV-A处理津田芜菁和赤丸芜菁块根24h后提取总RNA，通过RT-PCR方法克隆BrANS1和BrANS2基因并进行序列分析，通过Northern杂交检测BrANS1和BrANS2基因的表达。结果：BrANS1和BrANS2的开放读码框为1077bp，编码358个氨基酸残基；BRANS1和BRANS2与甘蓝ANS的同源性达97％，第211-307肽段具有20G-Fe（Ⅱ）加氧酶家族基因的结构域；BrANS1和BRANS2基因具有高度同源性，核苷酸序列在5个位置上存在差异，推导的氨基酸序列完全相同；uv-A可以诱导BrANS1和BrANS2表达，基因的表达量与处理时间相关。结论：克隆了津田芜菁和赤丸芜菁的BRANS1和BrANS2基因，这将为筛选依光型和非依光型花青素生物合成催化酶基因奠定研究基础。

Cloning,Sequence Analysis and Expression of Anthocyanidin Synthase Gene in Turnip

Objective： To clone the anthocyanidin synthase（ANS） genes of ‘Tsuda＇ and ‘Yurugi Akamaru＇ turnip, and to study the expression trait of the two ANS genes. Methods： The roots of ‘Tsuda＇ and ‘Yurugi Akamaru＇ turnip were irradiated with UV-A light for 24h. Total RNA was isolated and then BRANS1 and BRANS2 genes were cloned by RT-PCR method. Sequence analysis and the expression of BRANS1 and BRANS2 genes were conducted by bioinformatics method and Northern blotting. Results： The open reading frame（ORF） of BRANS1 and BRANS2 genes contained 1077 bp encoding proteins of 358 amino acids. Amino acid sequence analysis showed that BRANS1 and BRANS2 were 97% identitied to ANS of Brassica oleracea, and the 20G-Fe（Ⅱ） domain was in the sequence from 211 to 307. BRANS1 and BRANS2 genes had high identity. The nucleotide sequence of BRANS1 and BRANS2 genes had 5 differences and there were no difference in the deduced amino acid sequence. The expression of BRANS1 and BRANS2 could be induced by irradiation of UV-A, and the expression amount of the genes was correlated with light-exposure time. Conclusion： BRANS1 and BRANS2 genes of ‘Tsuda＇ and ＇Yurugi Akamaru＇ turnip were cloned successfully, which will establish the foundation for screening the catalyze enzyme genes involved in the light-sensitive and light-insensitve anthocyanidin biosynthesis.