解硫胺类芽孢杆菌SY20实时荧光定量PCR内参基因筛选
Screening of Reference Genes in Paenibacillus thiaminolyticus SY20 by Real-Time Fluorescene Quantitaticve PCR
吴雅萍 1杨技欣 2刘冬梅2
作者信息
- 1. 漳州职业技术学院食品工程学院,福建漳州 363000;华南理工大学食品科学与工程学院,广东广州 510640
- 2. 华南理工大学食品科学与工程学院,广东广州 510640
- 折叠
摘要
为研究解硫胺类芽孢杆菌SY20产多黏菌素相关基因的表达水平,需使用合适的内参基因进行荧光定量聚合酶链式反应法(Polymerase chain reaction,PCR)校正才能获得可信的检测结果.以RecA、RpoB、ftsZ、GAPDH及16S rRNA基因为候选基因进行内参基因筛选,利用荧光定量PCR分析这5个候选基因在解硫胺类芽孢杆菌SY20不同发酵时期的表达情况,再利用GeNorm、NormFinder、BestKeeper、比较Delta-Ct以及RefFinder 5个软件综合分析评估了5个候选内参基因的表达稳定性.结果显示,GeNorm分析表明ftsZ和RecA基因表达较为稳定,而NormFinder、BestKeeper、比较Delta-Ct及RefFinder分析表明ftsZ基因更为稳定.因此,结合5种分析方法确定ftsZ为最适合用于分析解硫胺类芽孢杆菌SY20基因表达的内参基因.
Abstract
In order to study the expression levels of polymyxin synthesis genes in Paenibacillus thiaminolyticus SY20,the appropriate reference gene is required to obtain reliable results in real-time quantitative polymerase chain reaction(PCR).The expressions of five candidate reference genes(RecA,RpoB,ftsZ,GAPDH and 16S rRNA)under different fermentation phases of Paenibacillus thiaminolyticus SY20 were analyzed by real-time fluorescence quantitative PCR,and their expression stabilities were evaluated by a comprehensive analysis of five software,including GeNorm,NormFinder,BestKeeper,the comparative Delta-Ct and RefFinder.The results showed that the genes of ftsZ and RecA were more stable using GeNorm analysis.But NormFinder,BestKeeper,comparative Delta-Ct and RefFinder analysis showed that ftsZ gene was the most stable.Therefore,the ftsZ gene was determined to be the most suitable reference gene for studying the gene expression of P.thiaminolyticus SY20 gene by combining five analytical methods.
关键词
解硫胺类芽孢杆菌/实时荧光定量PCR/内参基因筛选Key words
Paenibacillus thiaminolyticus/real-time fluorescence quantitative PCR/reference gene screening引用本文复制引用
基金项目
广东省自然科学基金(2021A1515012451)
漳州市食品产业技术研究院开放课题(ZSY2021114)
出版年
2024
NETL
NSTL
万方数据