title: Evaluation of Appropriate Reference Genes for Gene Expression Normalization during Watermelon Fruit Development.
abstract: Gene expression analysis in watermelon (Citrullus lanatus) fruit has drawn considerable attention with the availability of genome sequences to understand the regulatory mechanism of fruit development and to improve its quality. Real-time quantitative reverse-transcription PCR (qRT-PCR) is a routine technique for gene expression analysis. However, appropriate reference genes for transcript normalization in watermelon fruits have not been well characterized. The aim of this study was to evaluate the appropriateness of 12 genes for their potential use as reference genes in watermelon fruits. Expression variations of these genes were measured in 48 samples obtained from 12 successive developmental stages of parthenocarpic and fertilized fruits of two watermelon genotypes by using qRT-PCR analysis. Considering the effects of genotype, fruit setting method, and developmental stage, geNorm determined clathrin adaptor complex subunit (ClCAC), β-actin (ClACT), and alpha tubulin 5 (ClTUA5) as the multiple reference genes in watermelon fruit. Furthermore, ClCAC alone or together with SAND family protein (ClSAND) was ranked as the single or two best reference genes by NormFinder. By using the top-ranked reference genes to normalize the transcript abundance of phytoene synthase (ClPSY1), a good correlation between lycopene accumulation and ClPSY1 expression pattern was observed in ripening watermelon fruit. These validated reference genes will facilitate the accurate measurement of gene expression in the studies on watermelon fruit biology.
authors.affiliation: Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, China.
authors.forename: Qiusheng
authors.initials: Q
authors.lastname: Kong
authors.affiliation: Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, China.
authors.forename: Jingxian
authors.initials: J
authors.lastname: Yuan
authors.affiliation: Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, China.
authors.forename: Lingyun
authors.initials: L
authors.lastname: Gao
authors.affiliation: Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, China.
authors.forename: Liqiang
authors.initials: L
authors.lastname: Zhao
authors.affiliation: Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, China.
authors.forename: Fei
authors.initials: F
authors.lastname: Cheng
authors.affiliation: Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Huang
authors.affiliation: Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, China.
authors.forename: Zhilong
authors.initials: Z
authors.lastname: Bie
mesh_terms: D029773:Citrullus / Q000235:genetics* / Q000254:growth & development; D005638:Fruit / Q000235:genetics* / Q000254:growth & development; D020869:Gene Expression Profiling / Q000379:methods*; D018506:Gene Expression Regulation, Plant; D020043:Genes, Essential; D017343:Genes, Plant; D060888:Real-Time Polymerase Chain Reaction