《植物生理学报》 2021, 57(1): 195-204

铜胁迫下紫鸭跖草根组织实时定量PCR内参基因的选择

杨坤¹，黄超²，卢山¹，彭国颖¹，黄长干^1,3,*

¹江西农业大学理学院, 南昌330045；²江西农业大学校友办, 南昌330045；³南昌市植物资源化学利用重点实验室, 南昌330045

通信作者：黄长干；E-mail: windhcg@163.com

摘　要：

利用实时定量PCR技术分析不同铜浓度胁迫下紫鸭跖草(Setcreasea purpurea)根系的PCR标准化内参基因。从紫鸭跖草根系转录组数据库选取8个候选内参基因, 运用BestKeeper、Normfinder和GeNorm三款软件分析它们在不同铜浓度胁迫条件下紫鸭跖草根系的表达稳定性, 并利用2个目的基因MT和PCS对筛选出的内参基因组合进行验证。结果表明, 最佳的内参基因组合为18SrRNA、TUB和UBI; 2个目的基因MT和PCS在铜胁迫下的表达模式验证内参基因组合具有可靠性, 表明使用不合适的参考基因作为内控会导致结果不显著。

关键词：紫鸭跖草; 内参基因; 铜胁迫; 实时定量PCR

收稿：2019-10-14   修定：2020-11-17

资助：国家自然科学基金(31760068)。

Reference gene selection for quantitative real-time PCR in purple setcreasea (Setcreasea purpurea) root tissue under copper stress

YANG Kun¹, HUANG Chao², LU Shan¹, PENG Guoying¹, HUANG Changgan^1,3,*

¹College of Science, Jiangxi Agricultural University, Nanchang 330045, China; ²Alumni, Jiangxi Agricultural University, Nanchang 330045, China; ³Nanchang Key Laboratory of Chemical Utilization of Plant Resources, Nanchang 330045, China

Corresponding author: HUANG Changgan; E-mail: windhcg@163.com

Abstract:

Quantitative real-time PCR was used to analyze the PCR standardized reference genes in roots of Setcreasea purpurea under different concentrations of copper stresses. Eight candidate reference genes were selected from the transcriptome database of the root system of S. purpurea, and their expression stability under different concentrations of copper stresses were analyzed by using three softwares of Best-Keeper, NormFinder and GeNorm. The selected combination of reference genes was verified by using two target genes MT and PCS. The results showed that the best combination of reference genes was 18SrRNA, TUB and UBI. The expression patterns of two target genes MT and PCS under copper stress verified the reliability of the reference genes combination, indicating that the use of inappropriate reference genes as internal control would lead to insignificant results.

Key words: Setcreasea purpurea; reference genes; copper stress; quantitative real-time PCR