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Journal of Integrative Agriculture  2017, Vol. 16 Issue (04): 809-819    DOI: 10.1016/S2095-3119(16)61436-3
Crop Genetics · Breeding · Germplasm Resources Advanced Online Publication | Current Issue | Archive | Adv Search |
Identification of suitable reference genes in leaves and roots of rapeseed (Brassica napus L.) under different nutrient deficiencies
HAN Pei-pei*, QIN Lu*, LI Yin-shui, LIAO Xiang-sheng, XU Zi-xian, HU Xiao-jia, XIE Li-hua, YU Chang-bing, WU Yan-feng, LIAO Xing

Oil Crops Research Institute, Chinese Academy of Agricultural Sciences/Key Laboratory of Biology and Genetics Improvement of Oil Crops, Ministry of Agriculture, Wuhan 430062, P.R.China

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Abstract  Nutrient deficiency stresses often occur simultaneously in soil.  Thus, it’s necessary to investigate the mechanisms underlying plant responses to multiple stresses through identification of some key stress-responsive genes.  Quantitative real-time PCR (qRT-PCR) is essential for detecting the expression of the interested genes, of which the selection of suitable reference genes is a crucial step before qRT-PCR.  To date, reliable reference genes to normalize qRT-PCR data under different nutrient deficiencies have not been reported in plants.  In this study, expression of ten candidate reference genes was detected in leaves and roots of rapeseed (Brassica napus L.) after implementing different nutrient deficiencies for 14 days.  These candidate genes, included two traditionally used reference genes and eight genes selected from an RNA-Seq dataset.  Two software packages (GeNorm, NormFinder) were employed to evaluate candidate gene stability.  Results showed that VHA-E1 was the highest-ranked gene in leaves of nutrient-deficient rapeseed, while VHA-G1 and UBC21 were most stable in nutrient-deficient roots.  When rapeseed leaves and roots were combined, UBC21, HTB1, VHA-G1 and ACT7 were most stable among all samples.  To evaluate the stabilities of the highest-ranked genes, the relative expression of two target genes, BnTrx1;1 and BnPht1;3 were further determined.  The results showed that the relative expression of BnTrx1;1 depended on reference gene selection, suggesting that it’s necessary to evaluate the stability of reference gene prior to qRT-PCR.  This study provides suitable reference genes for gene expression analysis of rapeseed responses to different nutrient deficiencies, which is essential for elucidation of mechanisms underlying rapeseed responses to multiple nutrient deficiency stresses.
Keywords:   reference genes      rapeseed (Brassica napus L.)      nutrient deficiency      leaves      roots  
Received: 26 April 2016   Accepted:

This work was supported by the grants from the Agricultural Science and Technology Innovation Program, Chinese Academy of Agricultural Sciences (CAAS-ASTIP-2013-OCRI) and the Excellent Young Scientist Fund of Chinese Academy of Agricultural Sciences (1610172015004), and an open project funded by State Key Laboratory for the Conservation and Utilization of Subtropical Agro-bioresources, China (SKLCUSA-b201403).

Corresponding Authors:  LIAO Xing, Tel/Fax: +86-27-86819709, E-mail:    

Cite this article: 

HAN Pei-pei, QIN Lu, LI Yin-shui, LIAO Xiang-sheng, XU Zi-xian, HU Xiao-jia, XIE Li-hua, YU Chang-bing, WU Yan-feng, LIAO Xing. 2017. Identification of suitable reference genes in leaves and roots of rapeseed (Brassica napus L.) under different nutrient deficiencies. Journal of Integrative Agriculture, 16(04): 809-819.

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