掌叶大黄4个MYB转录因子的分子克隆及胁迫表达  

Molecular Cloning and Stress Expression of Four MYB Transcription Factors in Rheum palmatum

作　　者：赵霞 李元敏 李依民 胡晓晨 高静 颜永刚[1,2] 张岗 ZHAO Xia;LI Yuanmin;LI Yimin;HU Xiaochen;GAO Jing;YAN Yonggang;ZHANG Gang(College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine,Shaanxi University of Chinese Medicine,Xi’an 712046,China;Key Laboratory for Research and Development of“Qin Medicine”of Shaanxi Administration of Traditional Chinese Medicine,Shaanxi University of Chinese Medicine,Xi’an 712046,China;The Second Affiliated Hospital of Shaanxi University of Chinese Medicine,Xi’an 712046,China)

机构地区：[1]陕西中医药大学药学院/陕西省秦岭中草药应用开发工程技术研究中心,西安712046 [2]陕西中医药大学,陕西省中医药管理局“秦药”研发重点实验室,西安712046 [3]陕西中医药大学第二附属医院,西安712046

出　　处：《西北农业学报》2024年第7期1352-1363,共12页Acta Agriculturae Boreali-occidentalis Sinica

基　　金：国家自然科学基金(82104334,81973430);咸阳市中青年科技创新领军人才(L2022CXNLRC009);陕西中医药大学学科创新团队项目(2019-QN01)。

摘　　要：旨在克隆获得 RpMYB2、 RpMYB3、 RpMYB5、 RpMYB6基因ORF,并对其理化特性、系统进化关系、不同组织表达、激素与非生物胁迫下的表达等展开分析。研究结果表明, RpMYB2、 RpMYB3、 RpMYB5、 RpMYB6依次编码333、275、293、291个氨基酸,分子质量分别为34.781、30.525、31.243、33.313 ku, 4个RpMYBs蛋白均为亲水性蛋白并具有较高的热稳定性,除 RpMYB2外其余3个基因结构主要为α-螺旋和无规则卷曲,且均定位于细胞核。由蛋白结构域和保守基序分析发现, RpMYB2、 RpMYB5含有1个HTH-MYB结构域, RpMYB3、 RpMYB6含有2个HTH-MYB结构域,且不同MYB转录因子包含的保守元件数量及种类存在差异,系统进化分析显示 RpMYB2、 RpMYB5属于R1-MYB亚家族, RpMYB3、 RpMYB6属于R2R3-MYB亚家族,与其结构域分类一致。实时荧光定量结果显示,根中 RpMYB2、 RpMYB6转录本丰度最高, RpMYB3、 RpMYB5基因分别在叶片、叶柄中高表达。经200μmol·L^(-1)脱落酸(abscisic acid, ABA)处理, RpMYB5于24 h达峰值;200μmol·L^(-1)茉莉酸甲酯(methyl jasmonate, MeJA)诱导 RpMYB6表达,表达量随时间推移呈逐渐上升趋势且于24 h最高(为0 h的10.30倍),抑制 RpMYB3表达;200μmol·L^(-1)水杨酸(salicylic acid, SA)诱导 RpMYB6表达最为明显,3 h表达量上调至0 h的21.84倍。RpMYBs在非生物胁迫处理下表达也各有差异, RpMYB2受高温胁迫诱导表达, RpMYB3在损伤、高温胁迫下表达下调, RpMYB5在损伤胁迫下表达受抑制, RpMYB6响应高温、盐胁迫。In this study,we searched the transcriptome database to obtain the sequence information of four MYB gene family members,RpMYB2,RpMYB3,RpMYB5 and RpMYB6 genes.The corresponding physicochemical properties,phylogenetic relationships,tissue-specific expression,and induced expression under hormonal and abiotic stresses were analyzed.The results showed that RpMYB2,RpMYB3,RpMYB5 and RpMYB6 encoded 333,275,293 and 291 amino acids with molecular masses of 34.781,30.525,31.243 and 33.313 ku,respectively,and the four RpMYBs proteins were all hydrophilic proteins with high thermal stability,except for RpMYB2.The remaining three gene structures were mainlyα-helix and randomly coiled,and all of them were localized in the nucleus.Analysis of the protein structural domains and conserved motifs revealed that RpMYB2 and RpMYB5 contained one HTH-MYB structural domain,and RpMYB3 and RpMYB6 contained two HTH-MYB structural domains,and there were differences in the number and types of conserved elements contained in different MYB transcription factors.Phylogenetic analysis showed that RpMYB2 and RpMYB5 belonged to the R1-MYB subfamily,and RpMYB3 and RpMYB6 belonged to the R2R3-MYB subfamily,consistent with their structural domain classification.The protein structural domains and phylogenetic trees consistently indicated that RpMYB2 and RpMYB5 belonged to the R1-MYB subfamily,and RpMYB3 and RpMYB6 belonged to the R2R3-MYB subfamily.Real-time fluorescence quantification revealed that RpMYB2 and RpMYB6 were the most highly expressed in roots,and RpMYB3 and RpMYB5 were highly expressed in leaves and petioles.Respectively,RpMYB5 peaked at 24 h when treated with 200μmol·L^(-1)abscisic acid(ABA).The highest expression of RpMYB6 was induced by 200μmol·L^(-1)methyl jasmonate(MeJA),and the expression level increased gradually with time and was 10.30 times higher at 24 h than at 0 h,and inhibited RpMYB3 expression.200μmol·L^(-1)salicylic acid(SA)induced the most pronounced expression of RpMYB6 at 21.84 fold.The expression of RpMYBs also v

关 键 词：掌叶大黄 MYB 表达分析 胁迫 实时荧光定量PCR 

分 类 号：S567.239[农业科学—中草药栽培]