title: Selection and evaluation of novel reference genes for quantitative reverse transcription PCR (qRT-PCR) based on genome and transcriptome data in Brassica napus L.
abstract: Selection of reference genes in Brassica napus, a tetraploid (4×) species, is a very difficult task without information on genome and transcriptome. By now, only several traditional reference genes which show significant expression differentiation under different conditions are used in B. napus. In the present study, based on genome and transcriptome data of the rapeseed Zhongshuang-11 cultivar, 14 candidate reference genes were screened for investigation in different tissues, cultivars, and treated conditions of B. napus. These genes were as follows: ELF5, ENTH, F-BOX7, F-BOX2, FYPP1, GDI1, GYF, MCP2d, OTP80, PPR, SPOC, Unknown1, Unknown2 and UBA. Among them, excluding GYF and FYPP1, another 12 genes, were identified to perform better than traditional reference genes ACTIN7 and GAPDH. To further validate the accuracy of the newly developed reference genes in normalization, expression levels of BnCAT1 (B. napus catalase 1) in different rapeseed tissues and seedlings under stress conditions were normalized by the three most stable reference genes PPR, GDI1, and ENTH and little difference existed in normalization results. To the best of our knowledge, this is the first time B. napus reference genes have been provided with the help of complete genome and transcriptome information. The new reference genes provided in this study are more accurate than previously reported reference genes in quantifying expression levels of B. napus genes.
authors.affiliation: Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan 430062, PR China.
authors.forename: Hongli
authors.initials: H
authors.lastname: Yang
authors.affiliation: Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan 430062, PR China.
authors.forename: Jing
authors.initials: J
authors.lastname: Liu
authors.affiliation: Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan 430062, PR China.
authors.forename: Shunmou
authors.initials: S
authors.lastname: Huang
authors.affiliation: College of Life Sciences, Wuhan University, Wuhan 430072, PR China.
authors.forename: Tingting
authors.initials: T
authors.lastname: Guo
authors.affiliation: Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan 430062, PR China.
authors.forename: Linbin
authors.initials: L
authors.lastname: Deng
authors.affiliation: Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan 430062, PR China. Electronic address: huawei@oilcrops.cn.
authors.forename: Wei
authors.initials: W
authors.lastname: Hua
mesh_terms: D001937:Brassica / Q000235:genetics* / Q000378:metabolism; D017343:Genes, Plant; D005819:Genetic Markers; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000592:standards*; D059467:Transcriptome
chemical_list: D005819:Genetic Markers
keywords: B. napus; Normalization; Quantitative reverse transcription PCR; Reference gene; Stability; Transcriptome