根癌农杆菌胁迫下多花蔷薇(Rosamultiflora‘Inermis4’)的基因表达分析

Analysis of Genes of Rosa multiflora ‘Inermis4’ Expressed in Response to Agrobacterium tumefaciens Infection

多花蔷薇无刺4号(Rosamultiflora‘Inermis4’)对根癌病有较强抗性。以根癌农杆菌(Agrobacteriumtumefaciens)接种多花蔷薇无刺4号为目的材料,刺伤处理为对照,采用抑制差减杂交(suppressionsubtractivehybridization,SSH)方法,构建了富集多花蔷薇根癌病抗性相关基因的差减cDNA文库。差异筛选差减cDNA文库后,随机选取增强表达的92个克隆进行测序,去除冗余的cDNA和病菌的基因片段,共得到53个单一序列。利用GenBank数据库进行核酸和蛋白质同源性比较,获得的cDNA片段功能涉及:细胞分裂、生长(22个克隆),逆境防御、信号转导及激素调节(11个克隆)以及初生与次生代谢(10个克隆)。另有10个基因片段功能未知,可能为新的EST。根据测序及比对分析结果,对多花蔷薇无刺4号抗根癌病的分子机理进行了探讨。

Rosa multiflora ‘Inermis 4’ is a crown gall disease resistant ecotype. Suppression subtractive hybridization (SSH) was utilized to identify the host genes that were differentially expressed in the host-pathogen interactions. In vitro cultured shoots inoculated with Agrobacterium tumefaciens were used as “tester” and needle pricking shoots as “driver”. Subtractive cDNA library was constructed, which enriched for resistance-related genes to A. tumefaciens. Ninety-two cDNA clones were sequenced after differential screening through cDNA macroarray. After removing redundant and bacteria cDNA fragments, 53 unique cDNA clones were obtained. The putative function of these genes involved in:(1)cell division and growth and chaperones;(2)defense/stress, signaling, hormone regulation;(3)primary and secondary metabolism. Ten clones were designated as unknowns, these clones had either substantial similarity to genes with unknown functions, or only low similarity matches to GenBank database sequences. Furthermore, the probable molecular mechanism of R. multiflora ‘Inermis 4’ resistance to crown gall development was discussed according to the sequencing and blasting results.