甜樱桃花芽不同发育时期内参基因的筛选与验证

Selection and Validation of Reference Genes in Sweet Cherry Flower Bud at Different Development Stages

为了筛选甜樱桃花芽不同发育时期均稳定表达的内参基因,以甜樱桃桑提娜和黔樱一号不同发育时期花芽为材料,通过qRT-PCR技术检测28S rRNA、EF1-a1、EF1-a2、UBC、RPL13、18S rRNA、RSP3、CYP40、ACT2和α-TUB3等10个常用看家基因的表达水平,并利用GeNorm、NormFinder和BestKeeper综合评价其表达稳定性。结果表明,EF-1a2和RSP3在所有样品中稳定性最好。分别以EF-1a2、RSP3及EF-1a2+RSP3作为内参基因检测不同发育时期花芽生长素运输载体AUX1基因及生长素响应因子ARF基因的表达模式,该2个基因在不同内参基因标定下表达模式相同。表明EF-1a2、RSP3及EF-1a2+RSP3可作为甜樱桃花芽不同发育时期的内参基因。

In order to screen stably expressed reference genes in sweet cherry flower bud,the flower buds of sweet cherry verity Sangtina and Qianying 1 at different developmental stages were used as materials to investigate the expression levels of 10 house-keeping genes,including 28S rRNA,EF1-a1,EF1-a2,UBC,RPL13,18S rRNA,RSP3,CYP40,ACT2 andα-TUB3,based on qRT-PCR and the software of GeNorm,NormFinder and BestKeeper were also used to evaluate their expression stability.The results showed that EF1-a2 and RSP3 demonstrated the best stability among all samples.Additionally,the EF1-a2,RSP3 and EF1-a2+RSP3 were used as reference genes to detect the expression patterns of auxin transport-related gene AUX1 and auxin response-related gene ARF gene in flower buds at different stages of development and the results showed that there were the same expression patterns of the two genes under different calibration of internal reference genes.Simultaneously,the results also showed that EF1-a2,RSP3 and EF1-a2+RSP3 could be used as internal reference genes in sweet cherry flower buds at different developmental stages.