摘要
【目的】从橡胶树中克隆Glutathione-S-transferase(GST)基因的cDNA和基因组DNA,进行序列、结构和系统进化分析,研究胁迫条件和激素处理下的表达谱。【方法】利用产排胶机理课题组构建的胶乳EST数据库,通过PCR技术克隆橡胶树GST基因的cDNA和基因组DNA;在线预测蛋白质保守功能域和亚细胞定位,构建系统进化树,利用实时荧光定量PCR分析割胶、伤害、低温、激素和死皮病等因素对该基因表达的影响。【结果】从橡胶树胶乳中克隆到1个GST基因的全长cDNA(793 bp),编码25.4 kD(219aa)蛋白,命名为HbGSTU1;克隆了HbGSTU1的基因组DNA(1 313 bp),包含1个内含子(520 bp)和2个外显子。HbGSTU1属于GST Tau家族,推测是一种无信号肽的细胞质蛋白,其N端结构域包含结合GSH的G位点,C端结构域包含特异结合底物的H位点;HbGSTU1与一个蓖麻GST蛋白的亲缘关系最近,氨基酸序列的一致性为85%;HbGSTU1的表达受割胶、伤害、低温、2,4-D、乙烯利和死皮病等因素调控,但对JA、SA和ABA处理的应答不明显。【结论】HbGSTU1是一个典型的GST Tau家族蛋白,可能在橡胶树抗逆过程中起作用,可作为橡胶树抗逆分子育种的靶标基因。
【Objcetive】The objective of this study is to clone the cDNA and genomic clone of a glutathione-S-transferase（GST） from rubber tree,and characterize its structural and phylogenetic characteristics,investigate its expression patterns under various abiotic stresses and hormone treatments.【Method】Combined with the latex EST database,RT-PCR and PCR were used to clone the full length cDNA and genomic DNA of a Hevea GST,respectively.On-line bioinformatics tools were used to predict the conserved functional domains and subcellular localization.Clustal X was used to construct the phylogenetic tree.Real-time RT-PCR（qRT-PCR） was used to characterize the expression patterns of the Hevea GST gene under various treatments（tapping,wounding,chilling,hormone and tapping panel dryness）.【Result】The full-length cDNA（793 bp） encoding a 25.4 kD（219 aa） Tau GST protein was firstly cloned from the latex of H.brasiliensis,named HbGSTU1 and deposited in GenBank（JF729318）.The genomic DNA（1 313 bp） of HbGSTU1 was also cloned,which contains one intron（520 bp） and two exons.HbGSTU1 protein contained the Tau GST-specific N-terminal domain（G site） and the C-terminal domain（H site）.HbGSTU1 was predicted to be a cytoplasmic protein,which shared the highest amino acid identity（85%） with a GST from Ricinus communis.The qRT-PCR results show that HbGSTU1 expression was regulated by tapping,wounding,4℃ chilling,2,4-D and Ethrel,but not by the three hormones JA,SA and ABA.【Conclusion】HbGSTU1 is a typical Tau GST protein,and is implicated in stress-responses in Hevea tree,which may serve as a target gene for Hevea molecular breeding.
出处
《中国农业科学》
CAS
CSCD
北大核心
2011年第20期4150-4158,共9页
Scientia Agricultura Sinica
基金
国家"973"计划项目(2008CB117009)
天然橡胶产业技术体系育种岗
中国热带农业科学院橡胶研究所基本科研业务费专项资金(XJSYWFZX2010-05(N))
关键词
橡胶树
谷胱甘肽-S-转移酶
基因表达
胁迫
激素
Hevea brasiliensis
glutathione-S-transferases
gene expression
stress
hormones
