中国中药杂志2025，Vol.50Issue(3)：682-692,11.DOI:10.19540/j.cnki.cjcmm.20241113.101
土家族药血筒实时荧光定量PCR内参基因的筛选和验证
Selection and validation of reference genes for quantitative real-time PCR analysis in Tujia medicine Xuetong
摘要
Abstract
Tujia ethnic group medicine Xuetong is derived from Kadsura heteroclita,the stem of which has the medicinal value for anti-rheumatoid arthritis,liver protection,anti-tumor,anti-oxidation effects,and has been widely used in Hunan and Guangdong in China.The selection of reliable and stable reference genes is the basis for subsequent molecular research on K.heteroclita.In this study,GAPDH,TUA,Actin,UBQ,EF-1α,18S-rRNA,CYP,UBC,TUB,H2A,and RPL were selected as candidate reference genes in Kadsura heteroclita.The gene expression levels of the 11 candidate reference genes of K.heteroclita in its 6 different parts(stem-inside of the cambium,stem-outside of the cambium,fruit,flower,root,and leaf)and under different intervention conditions[drought stress,salt stress,and methyl jasmonate(MeJA)treatment]were detected by quantitative real-time polymerase chain reaction(qRT-PCR).The expression stability of the 11 candidate reference genes was comprehensively analyzed and evaluated by geNorm,NormFinder,ΔCT algorithm,and RefFinder software.The results showed that the expression of UBC and RPL was relatively stable in 6 different parts,and UBC and GAPDH genes were relatively stable under different intervention conditions.To verify the reliability of reference genes for K.heteroclita,this study further examined the relative expression levels of KhFPS,KhIDI,KhCAS,KhSQE,KhSQS,KhSQS-2,KhHMGS,KhHMGR,KhMVD,KhMVK,KhDXR,KhDXS,KhPMVK,and KhGGPS in different parts and under different intervention conditions,which might relate to the synthesis of the main component(Xuetongsu)of K.heteroclita.The results showed that with UBC and RPL or UBC and GAPDH as the reference genes,the expression trends of these 14 genes were basically consistent in different parts or under different intervention conditions for K.heteroclita.In conclusion,UBC can be used as a reference gene of K.heteroclita for its different parts and different intervention conditions,which lays a foundation for further research on the biosynthetic pathway of main components in K.heteroclita.关键词
异形南五味子/实时荧光定量PCR/内参基因/UBCKey words
Kadsura heteroclita/quantitative real-time PCR/reference gene/UBC引用本文复制引用
肖倩,谭琛偲,曾江,徐媛菽,符天昊,宁露云,王炜..土家族药血筒实时荧光定量PCR内参基因的筛选和验证[J].中国中药杂志,2025,50(3):682-692,11.基金项目
湖南省自然科学基金项目(2023JJ40482) （2023JJ40482）
湖南省教育厅科学研究项目(22C0189) （22C0189）
2022年度中医药民族医药国际联合实验室开放基金项目(2022GJSYS09) （2022GJSYS09）
湖南中医药大学本科生科研创新项目(2023BKS115,2024BKS079) （2023BKS115,2024BKS079）
