松材线虫体前端特异cDNA文库的构建及细胞壁松弛蛋白基因的克隆与初步分析

Construction of specific cDNA library and cloning of expansin gene from the ante-rior end of Bursaphelenchus xylophilus

本研究将松材线虫切割为包括食道腺、分泌孔和头感器的体前端以及包括肠道的体后端,通过改进的固相杂交方法进行消减杂交,构建了松材线虫体前端的特异cDNA文库,利用该文库并结合RACE技术获得了松材线虫细胞壁松弛蛋白基因的全长cDNA。该基因编码149个氨基酸,经过比对发现其翻译产物与马铃薯金线虫EXPB1、EXPB2蛋白有46%和42%的相似性,与南方根结线虫的推定无毒基因蛋白(CAC27774.1)有39%的相似性,该基因产物可能在侵染过程中起着软化寄主细胞壁的作用,但具体功能有待进一步研究。

For isolation and identification of parasitic genes of the nematode, pine wood nematode was cut into two parts, and a specific cDNA library of anterior end of nematode was constructed by modified solidphase subtractive hybridization. A full-length cDNA of putative expansin of pine wood nematode was obtained from the subtractive hybridized library with RACE （ rapid amplification of cDNA ends）. The cDNA designed as Bx0B14 contained an open reading frame encoding 149 amino acids and was 42%, 46% and 39% identity with EXPB1, EXPB2 from G. rostochiensis and putative avirulence protein （ CAC27774.1 ） from M. incognita. The expansin of pine wood nematode may have a role in loosing cell walls during invading its host plant, but its specific function is not clearly.