龙眼miR396分子进化特性及响应蓝光的表达分析
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引用本文:李汉生,孙 刚,陈晓慧,姚德恒,林玉玲,赖钟雄.龙眼miR396分子进化特性及响应蓝光的表达分析[J].西北植物学报,2019,39(12):2109~2122
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作者单位
李汉生1,2,孙 刚2,陈晓慧1,姚德恒1,林玉玲1,赖钟雄1* (1 福建农林大学 园艺植物生物工程研究所福州3500022 三明学院 资源与化工学院福建三明365004) 
基金项目:国家自然科学基金(31572088,31672127);
中文摘要:关于蓝光对龙眼胚性愈伤组织miRNA组学研究中发现大量响应蓝光的miRNAs,挑选关键且差异表达的miR396a 3p_2和miR396b 5p进行分子进化特性分析,以进一步明确它们在龙眼响应蓝光过程中的作用。该研究对前体和成熟体进化树、前体二级结构、启动子顺式作用元件、靶基因预测及其响应蓝光表达模式等进行分析。结果表明:(1)由5p臂上形成的miR396成熟体序列保守性较高,由3p臂上形成的miR396成熟体序列特异性较大;茎序列的保守性高于环序列,5p臂上保守性高于3p臂。(2)miR396a 3p_2和miR396b 5p的启动子主要包括光、激素、生物和非生物胁迫响应元件,可能通过这些顺式元件在龙眼响应蓝光中起调控作用;其响应蓝光的靶基因或蛋白主要包括生长素调控因子(GRF2)、LRR类受体丝氨酸(FLS2)、乙烯不敏感蛋白(EIN3)和DNA定向RNA聚合酶α亚基(rpoA)等。(3)实时荧光定量 PCR 结果表明,miR396a 3p_2和miR396b 5p在不同光质的表达模式存在差异; 在不同光强的蓝光条件下,miR396b 5p与其靶基因的表达趋势基本呈负相关,而miR396a 3p_2与rpoA的表达趋势未呈负相关,可能存在其他miR396家族成员或miRNAs同时调控,从而实现靶基因转录水平的调控。
中文关键词:龙眼  miR396  进化特性  蓝光  实时荧光定量PCR
 
Analysis on Evolutionary Characteristics and Expression Patterns of miR396 Responding to Blue Light in Dimocarpus longan
Abstract:Based on the analysis of miRNAs omics in longan ECs, we selected the differentially expressed miR396a 3p_2 and miR396b 5p for further analysis and verification, which play a key role in the regulatory pathway about effect of blue light on longan. We analyzed the phylogenetic tree of precursor and mature sequences, the secondary structure and cis acting element of precursor, the prediction of target genes, as well as expression patterns responding to blue light of miR396 in this study. These results showed that: (1) the miR396 mature sequences from 5p arm were highly conserved and from 3p arm were quite specific. The stem sequence was more conserved than loop sequence, and the miRNA from 5p arm sequence was more conserved than that from the 3p arm. (2) They both contain many light responsive elements, hormone responsive elements, and biotic and abiotic stress responsive elements, which might play a regulatory role in response to blue light through these cis elements. Their targets responding to blue light both contain growth regulating factor 2 (GRF2), LRR receptor like serine/threonine protein kinase (FLS2), ethylene insensitive protein 3 (EIN3) and DNA directed RNA polymerase subunit alpha (ropA) etc. Real time quantitative PCR showed that there were differences in the expression patterns of miR396a 3p_2 and miR396b 5p under different light qualities. The expression trends of miR396b 5p and its targets presented negative correlation under the blue light conditions with different light intensities, while miR396a 3p_2 was not negatively correlated with rpoA. This might be the reason that other miR396 family members or miRNAs were also involved in this process to achieve regulation of target gene transcription levels.
keywords:Dimocarpus longan  miR396  evolution  blue light  real time quantitative PCR
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