菜薹抽薹相关基因BrcuDFR-like/BrcuAXS的克隆与表达特性分析

Cloning and Expression Analysis of BrcuDFR-like/BrcuAXS Gene in Flowering Chinese Cabbage

为了预测抽薹相关基因BrcuDFR-like/BrcuAXS的功能,通过PCR和RACE的方法克隆了菜薹BrcuDFR-like/BrcuAXS基因的cDNA和gDNA全长序列。结果表明:该基因编码区全长1332bp,编码312个氨基酸残基。对应的gDNA全长为2460bp,含有8个外显子和7个内含子,内含子总长为1042bp,其中第3个内含子最长,为401bp。内含子中含有多个基本转录元件和顺式作用元件,如光应答元件、赤霉素响应元件、参与抗性和胁迫应答元件、热响应元件、WRKY转录因子的结合位点及干旱胁迫元件MYB转录因子结合位点等。利用半定量RT-PCR分析表达模式,发现BrcuDFR-like/BrcuAXS随菜薹花芽形态逐步建成直至抽薹开花,其表达量逐渐增强,与其它物种DFR-like基因的表达模式更吻合,由此预测该基因在菜薹生长发育阶段编码DFR-like酶的可能性大于编码AXS的可能性,其功能可能与菜薹营养分生组织向花分生组织转变有关。

In order to predict the function of BrcuDFR-like/BrcuAXS, both the full-length of cDNA and genomic DNA were cloned with the method of PCR and RACE and expression pattern was investigated in flowering Chinese cabbage （Brassiea rapa syn. campestris L. ssp. chinensis var. utilis Tsen et Lee） . The results indicated that the eDNA with the complete coding region was 1 332 bp in length which encoded 312 putative amino acids. The corresponding gDNA was 2 460 bp in length which harbored eight exons and seven introns. The longest intron was the third intron with 401 bp in length. A computer scan disclosed that the introns harbored light-responsive element, gibberellin-responsive element, defense and stress responsiveness element, heat stress responsiveness element, WRKY and MYB binding site and so on. The semi-quantitative RT-PCR analysis revealed that no detectable levels were expressed during the first stage of sampling, then transcripts were detected during the two true-leaf, the four true-leaf, the five true-leaf and the flowering stages. Based on expression analysis, it is more likely to encode DFR than AXS, and it may play a role of transition from vegetative growth to reproductive growth in flowering Chinese cabbage.