利用Illumina Hi SeqTM2500平台对光暗培养的杜仲愈伤组织进行了转录组高通量测序,利用BLAST软件进行差异表达基因的功能注释和富集分析,就白光(光强为12 000 lux,16 h光照,8 h黑暗)对杜仲愈伤组织中绿原酸含量的影响进行了研究.结果表明:通过Trinity软件合并组装后共获得62 030个Unigenes,通过BLASTX比对,共获得25 417(40.98%)个有注释信息的Unigenes.通过对KEGG通路进行深入分析表明,白光(光强为12 000 lux,16 h光照,8 h黑暗)培养杜仲愈伤组织18 d,与绿原酸合成相关的3种酶基因上调表达,它们分别为苯丙氨酸解氨酶(EC 4.3.1.24,phenylalanine ammonia-lyase,PAL)、肉桂酸-4-羟基化酶(EC 1.14.13.11,trans-cinnamate 4-monooxygenase,C4H)、奎宁酸羟基肉桂酰转移酶(EC 2.3.1.133,Shikimate o-hydroxyl-cinnamoyltransferase,HCT).由此推断,白光能促进杜仲愈伤组织中绿原酸的积累.研究结果为获取高产绿原酸资源和杜仲遗传分析提供有价值的资料.
In this study,Eucommia ulmoides calli for 18 d under white light( light intensity was 12 000 lux,16 h light and 8 h darkness) were cultured and 24 h darkness culture was as control. Then transcriptome high-throughput sequencing for the Eucommia ulmoides calli were performed using the Illumina Hi SeqTM2500 sequencing platform.Functional annotation and enrichment analysis of differentially expressed genes were also carried out. Effect of white light on the content of chlorogenic acid of Eucommia ulmoides calli was studied. The results showed de novo assembly generated 62 030 unigenes. 25 417( 40. 98%) annotated unigenes were obtained by selecting BLAST parameter( E-value ≤10^-5) and HMMER parameter( E-value ≤10^-10). Metabolic pathway analysis revealed that 3 unigenes were predicted to be responsible for the chlorogenic acid,the 3 unigenes were up-regulated and in the calli under light culture,which was meaning that the production capacity of chlorogenic acid increased. The three enzymes dominated chlorogenic acid production in Eucommia ulmoides calli. The three enzymes were phenylalanine ammonia-lyase( EC 4. 3. 1. 24,PAL),trans-cinnamate 4-monooxygenase( EC 1. 14. 13. 11,C4H) and Shikimate o-hydroxyl-cinnamoyltransferase( EC 2. 3. 1. 133,HCT). The study showed that the white light could improve the production capacity of chlorogenic acid components of Eucommia ulmoides calli and represents a valuable resource for future genomic studies on Eucommia ulmoides.