大豆Pre-miRNAs作为内参基因在盐碱胁迫下的表达稳定性分析  

Stability of Pre-miRNAs in soybean as reference genes for quantitative polymerase chain reaction under salt and alkali stresses

作　　者：周永刚[1,2] 王骐[3] 刘伟灿[1,2] 邓宇[1,2] 李晰亮 靳京[1,2] 邓文波[1,2] 赵利旦 王兴超[1,2] 王南[1,2] 王法微[1,2] 李晓薇[1,2] 董园园[1,2] 李海燕[1,2] 

机构地区：[1]吉林农业大学生物反应器与药物开发教育部工程研究中心,吉林长春130118 [2]吉林农业大学生命科学学院,吉林长春130118 [3]东北师范大学附属中学,吉林长春130118

出　　处：《西北农林科技大学学报（自然科学版）》2016年第1期61-67,共7页Journal of Northwest A&F University(Natural Science Edition)

基　　金：国家转基因生物新品种培育重大专项(2014ZX08010-002);国家自然科学基金项目(31271746;31201144);吉林省发改委项目(JF2012C002-04)

摘　　要：【目的】对比大豆Pre-miRNAs候选内参基因和传统内参基因的表达稳定性,筛选出适合盐、碱胁迫条件下RT-qPCR分析的最稳定内参基因。【方法】选用了6个保守的Pre-miRNAs基因和4个常规的内参基因作为候选内参基因,通过RT-qPCR的方法,利用GeNorm和NormFinder软件,评价了10个候选内参基因在大豆盐、碱胁迫条件下的稳定性。【结果】大豆盐胁迫下,叶中表达最稳定的基因组合是Pre-miR166和Pre-miR172,表达最稳定的单一基因是FboX;根中表达最稳定的基因组合是Act11和EF1A,表达最稳定的单一基因是Act11。大豆碱胁迫下,叶中表达最稳定的基因组合是Pre-miR393和Pre-miR172,表达最稳定的单一基因是Pre-miR393;根中表达最稳定的基因组合是Act11和60S,表达最稳定的单一基因是Pre-miR172。【结论】大豆Pre-miRNAs可以与传统内参基因一样作为内参定量目的基因。[Objective] This study selected the appropriate reference genes for stability of candidate precursor miRNAs and traditional housekeeping genes in bean under different stress treatments. [Method] Six conservative Pre miRNAs RT-qPCR by comparing different tissues of soy genes and four regularhousekeeping genes in soybean were selected as candidate reference genes to evaluate their expression sta bility under salt and alkali stresses during RT-qPCR process using GeNorm and NormFinder. [Result] In leaves,the most stable combination of genes under salt stress was Pre-miR166 and Pre-miR172 whereas the best single gene under this stress was FboX. While, in roots, the most stable combination of genes under salt stress was Act11 and EF1A whereas the best single gene was Act11. Under alkali stress conditions,the most stable combination of genes in leaves was Pre-miR89g and Pre-miR172 whereas the best single gene was Pre-miR393. The most stable combination of genes in roots was Actll and 60S whereas the best single gene was Pre-miR172. [Conclusion] Precursor miRNAs reference genes can be used as reference genes to gether with traditional housekeeping genes.

关 键 词：大豆 盐胁迫 碱胁迫 荧光定量PCR Pre-miRNAs(前体miRNAs) 内参基因 

分 类 号：Q-03[生物学] S-3[农业科学]