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以番茄(Lycopersicon esculentum)紫色花青素积累品系Aft基因型‘LA1996’为试验材料,以番茄总RNA反转录的c DNA文库为模板,根据mirbase数据库中MIR390基因和番茄unigene数据库预测到的靶基因序列,克隆番茄Lemi R390前体基因及其预测靶基因Le TAS3;采用定量PCR技术检测Lemi R390及其预测靶基因Le TAS3在番茄果实不同发育阶段的表达情况,利用RLM 5′RACE技术验证Lemi R390对靶基因m RNA的剪切作用。结果表明,克隆到两个Lemi R390的前体,分别与MIR390a和MIR390b一致,含有完整的茎环结构。Lemi R390与其靶基因Le TAS3在番茄果实不同发育时期表达量不同,果实发育前期特别是花后1~2周幼果期表达量高,发育后期均下降为微量表达,并且在幼果期Lemi R390与Le TAS3表达呈负调控关系。Lemi R390能够靶作用于Le TAS3的转录本调控其降解,其剪切位点为Lemi R390序列5′端的第10位碱基。
The Aft genotype ’LA1996’ of Lycopersicon esculentum accumulating line was used as the experimental material, and the reverse transcribed c DNA library of tomato total RNA was used as a template. According to the predicted target of MIR390 gene and tomato unigene database in mirbase database The Lemi R390 precursor gene and its predicted target gene Le TAS3 were cloned and sequenced. The expression of Lemi R390 and its predicted target gene Le TAS3 in different developmental stages of tomato fruit were detected by quantitative PCR. The expression of Lemi R390 on the target gene m RNA shearing effect. The results showed that two precursors cloned into Lemi R390 were identical to MIR390a and MIR390b, respectively, and contained a complete stem-loop structure. Lemi R390 and its target gene Le TAS3 expressed differently in tomato fruit at different developmental stages. The early stage of fruit development, especially in young fruit 1 to 2 weeks after anthesis, was significantly decreased in young fruit stage, and decreased in young fruit stage. Lemi R390 and Le TAS3 expression was negatively regulated. Lemi R390 is able to target the transcripts of Le TAS3 to regulate its degradation at its cleavage site at base 10 of the 5 ’end of the Lemi R390 sequence.