杨树受溃疡病菌感染后转录因子的应答变化  

Response of Transcription Factors of Populus tomentosa to Inoculation with Botryosphaeria dothidea

作　　者：张秀英[1] 宋瑞清[2] 张星耀[3] 

机构地区：[1]长春工业大学化学与生命科学学院,长春130000 [2]东北林业大学林学院,哈尔滨150040 [3]中国林业科学研究院森林生态环境与保护研究所,北京100091

出　　处：《林业科学》2015年第4期110-115,共6页Scientia Silvae Sinicae

基　　金：林业公益性行业科研专项"重大森林病虫灾害防控技术的关键理论基础"(201204501);中国林业科学研究院院部基金"林木病害和病原标本及病原体的收藏保存和评价"(CAFYBB2011005-4);科技部基础性工作专项"中国树木溃疡病原多样性及其生态地理分布和危害调查"(2009FY210100)

摘　　要：【目的】研究杨树受溃疡病菌感染后转录因子的应答变化。【方法】提取诱导前后的毛白杨树皮mRNA，反转录生成 cDNA，构建接种溃疡病菌72 h 后的 SSH － cDNA 库，从库中获得 ESTs，将 ESTs 注释到Unisequences，挑选出与转录相关的Unisequences，利用RT－qPCR对转录基因作进一步的分析。随机挑选199个阳性克隆进行测序，测序结果经与 NCBI基因库中序列进行比较，发现172个 EST 同源序列，对这些 EST 在 dbEST 数据库进行同源检索分析。【结果】有10个序列与转录相关，占5．8%，分为4类，即：锌指蛋白，含EAR转录区域；类CONSTANS转录因子，编码核蛋白；普通转录因子，调节转录；WRKY21转录因子，参与防御信号转录。通过 RT －qPCR对其中3类（锌指蛋白、类 CONSTANS和 WRKY21）转录因子做进一步分析发现，Z锌指蛋白（ Cys/His－rich）在接种溃疡病菌后，在诱导初期增长缓慢，48 h后逐渐升高，144 h达到最高，表达量是未接种时的50倍；WRKY21表达量先升高（12～24 h），然后降低（24～96 h），144 h 时，表达量是未接种时0.94倍；类 CONSTANS 表达量一直处于缓慢升高的趋势，直到诱导144 h，是未接种时2倍。【结论】毛白杨被溃疡病菌感染后与防御相关的转录因子表达量明显上升，这可为毛白杨抗溃疡病重要功能基因的克隆和鉴定提供理论基础。[Objective]This study aims at investigating the response of transcription-related genes of Populus tomentosa induced by Botryosphaeria dothidea infectation.[Method]The suppression subtractive hybridization （ SSH） technique was used to construct a cDNA library,and the bioinformatics were used to analyze the expression of transcription related gene of Populus tomentosa infected with Botryosphaeria dothidea. The bark mRNA was purified and transcribed into cDNA. A poplar-pathogen interaction cDNA library of inoculated poplar bark 72 h post-inoculation （ hpi） was constructed. Expressed sequence tags （ ESTs） were obtained,which were assembled into unisequences prior to their annotation. The unisequences related to transcription were selected,and were further analyzed by quantitative real-time PCR （ qRT-PCR） . The randomly selected 199 positive clones were sequenced,and compared with the sequences of NCBI gene pool,among which 172 EST sequences were found homologous. The homologous analysis for these EST was conducted in dbEST database retrieval,[Result]The result showed that 10 sequences were associated with transcription,and accounted for 5. 8% of the total sequences. They were divided into four categories,i. e. Zinc finger protein （ Cys / His-rich）,containing the EAR transcriptional repressor; CONSTANS-like protein,Encoding a nuclear protein; ordinary transcription factor,regulating transcription and WRKY21 transcription factor,participating in the plant defense. Through further RT-qPCR analyses on three kinds of transcription factors,the result showed that the expression amount of Zinc finger protein （ Cys / His-rich ） after inoculation was 50 times of the non-inoculated;The WRKY21 expression amount was 0. 94 times of the non-inoculated;The CONSTANS-like protein expression amount was 2 times of the non-inoculated.[Conclusion]The expression of these transcription factor genes was significantly increased in Populus tomentosa infected by Botryosphaeria dothidea.

关 键 词：葡萄座腔菌 毛白杨 抑制消减杂交 实时定量PCR(qRT—PCR) 转录因子 

分 类 号：S763.11[农业科学—森林保护学]