Validation of Reference Genes for Gene Expression by Quantitative Real-Time RT-PCR in Stem Segments Spanning Primary to Secondary Growth in Populus tomentosa.
The vertical segments of Populus stems are an ideal experimental system for analyzing the gene expression patterns involved in primary and secondary growth during wood formation. Suitable internal control genes are indispensable to quantitative real time PCR (qRT-PCR) assays of gene expression. In this study, the expression stability of eight candidate reference genes was evaluated in a series of vertical stem segments of Populus tomentosa. Analysis through software packages geNorm, NormFinder and BestKeeper showed that genes ribosomal protein (RP) and tubulin beta (TUBB) were the most unstable across the developmental stages of P. tomentosa stems, and the combination of the three reference genes, eukaryotic translation initiation factor 5A (eIF5A), Actin (ACT6) and elongation factor 1-beta (EF1-beta) can provide accurate and reliable normalization of qRT-PCR analysis for target gene expression in stem segments undergoing primary and secondary growth in P. tomentosa. These results provide crucial information for transcriptional analysis in the P. tomentosa stem, which may help to improve the quality of gene expression data in these vertical stem segments, which constitute an excellent plant system for the study of wood formation.
Beijing Key Laboratory of Agriculture Gene Resource and Biotechnology, Beijing Academy of Agriculture and Forestry Sciences, Beijing, P. R. China.
Ying
Y
Wang
Beijing Key Laboratory of Agriculture Gene Resource and Biotechnology, Beijing Academy of Agriculture and Forestry Sciences, Beijing, P. R. China.
Yajuan
Y
Chen
Beijing Key Laboratory of Agriculture Gene Resource and Biotechnology, Beijing Academy of Agriculture and Forestry Sciences, Beijing, P. R. China.
Liping
L
Ding
Beijing Key Laboratory of Agriculture Gene Resource and Biotechnology, Beijing Academy of Agriculture and Forestry Sciences, Beijing, P. R. China.
Jiewei
J
Zhang
Beijing Key Laboratory of Agriculture Gene Resource and Biotechnology, Beijing Academy of Agriculture and Forestry Sciences, Beijing, P. R. China.
Jianhua
J
Wei
Beijing Key Laboratory of Agriculture Gene Resource and Biotechnology, Beijing Academy of Agriculture and Forestry Sciences, Beijing, P. R. China.
Hongzhi
http://orcid.org/0000-0001-9052-9319
H
Wang
D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D018547:Plant Stems / Q000235:genetics* / Q000254:growth & development*; D032107:Populus / Q000235:genetics* / Q000254:growth & development*; D018749:RNA, Plant / Q000235:genetics / Q000302:isolation & purification; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D014158:Transcription, Genetic
D018749:RNA, Plant