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[1]蒋梦琦 苏立遥 黄倏祺 陈旭 徐小萍 张梓浩 赖钟雄 林玉玲**.龙眼miR156家族及其调控靶标SPL的生物信息学和表达模式*[J].应用与环境生物学报,2020,26(03):1-12.[doi:10.19675/j.cnki.1006-687x.2019.07024]
 JIANG Mengqi,SU Liyao,HUANG Shuqi,et al.Bioinformatics and expression pattern analysis of miR156 family and its regulatory target SPL in Dimocarpus longan*[J].Chinese Journal of Applied & Environmental Biology,2020,26(03):1-12.[doi:10.19675/j.cnki.1006-687x.2019.07024]
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龙眼miR156家族及其调控靶标SPL的生物信息学和表达模式*()
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《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
26卷
期数:
2020年03期
页码:
1-12
栏目:
研究论文
出版日期:
2020-06-25

文章信息/Info

Title:
Bioinformatics and expression pattern analysis of miR156 family and its regulatory target SPL in Dimocarpus longan*
作者:
蒋梦琦 苏立遥 黄倏祺 陈旭 徐小萍 张梓浩 赖钟雄 林玉玲**
福建农林大学园艺植物生物工程研究所 福州 350002
Author(s):
JIANG Mengqi;? SU Liyao;? HUANG Shuqi;? CHEN Xu;? XU Xiaoping;? Zhang Zihao;?LAI Zhongxiong & LIN Yuling**
Institute of Horticultural Biotechnology, Fujian Agriculture and Forestry University, Fuzhou 350002, China
关键词:
龙眼;?miR156;?进化特性;?体胚发生;?SPL
Keywords:
Dimocarpus longan;?MiR156;?Evolutionary characteristics;?Somatic embryogenesis;?SPL
DOI:
10.19675/j.cnki.1006-687x.2019.07024
摘要:
为了解miR156基因家族的进化特性及其在龙眼早期体胚发生过程中的调控作用,对植物miR156家族成员的成熟体和前体序列进行多重比对和进化特性分析,并对龙眼miR156基因家族成员进行了前体(pre-miRNA)二级结构预测、靶基因预测及裂解位点验证,利用qPCR技术探究龙眼体胚发生早期miR156a与靶基因SPL6/9/16调控模式。结果表明:龙眼miR156家族含有13条成熟体序列和6条前体序列。Mfold 预测显示pre-miR156家族6个成员的序列均能形成典型稳定的茎环二级结构,它们的最小折叠自由能(ΔG)在 - 43.30kal·mol-1 ~ -62.60kal·mol-1;成熟序列的碱基保守性高。系统发育进化树分析显示,龙眼miR156家族与水稻、拟南芥的miR156亲缘关系更为接近。靶基因预测显示,龙眼miR156a的靶基因是SPL2、SPL6、SPL9、SPL16、SPL18;利用改良的RLM-RACE在龙眼愈伤组织中检测到SPL6/9/16的断裂片段,miR156a在与靶基因mRNA互补的第10个核苷酸处切割靶标mRNA。实时荧光定量PCR显示,在龙眼体胚发生早期miR156a在第0-9天表达平稳,仅在球形胚(GE,globular embryos)阶段显着下调,SPL6/9/16表达量持续上调,在GE阶段表达量最高,提示SPL6/9/16的大量积累对龙眼球形胚形态建成具有重要作用。综上所述,植物miR156基因家族成熟体序列高度保守,且miR156a的下调表达与靶基因SPL6/9/16的大量积累可能对龙眼球形胚的建成具有重要意义。
Abstract:
MiR156 gene family is one of the ancient and conservative families and it involved in the regulation of plant growth and development. However, the miR156 gene family evolutionary characteristics and its function in Dimocarpus longan somatic embryos have not been reported yet. Therefore we investigated the evolution characteristics of miR156 gene family and its regulatory role in the early somatic embryogenesis of longan. we analyzed the mature and precursor sequences, and the phylogenetic of longon and other plants , predicted the secondary structure of precursors miR156 (pre-miR156) family members , analyzed the regulatory patterns of miR156a and its target genes SPL6/9/16 during the early somatic embryogenesis of longan. The longan miR156 family contain ed 13 mature sequences and 6 precursor sequences. Mfold prediction showed that the sequences of the pre-mir156 family members could form a typical stable secondary stem-loop structures , and their minimum folding free energy (ΔG) ranged from -43.30 kal·mol-1 to - 62.60kal·mol-1 ; Multiple sequences alignment of these pre-miR1 56 members showed that the regions generating mature miRNA were highly conservative. Phylogenetic tree analysis showed that miR156 family of longan was more closely related to those from rice and Arabidopsis thaliana. According to prediction of target gene, the target genes of miR156a were SPL2, SPL6, SPL9, SPL16 and SPL18. We detected SPL6/9/16 fragments in longan callus using an improved RLM-RACE, and miR156a cleaved the target mRNA at the 10th nucleotide. Real-time fluorescence quantitative PCR showed that in the early somatic embryos of longan, the expression of miR156a was stable at day 0-9th, but only decreased significantly in globular embryos (GE) , while SPL6/9/16 was increased significantly. These results showed that the mature sequences of miR156 families in plants are highly conserved , the decreased miR156 and the accumulation of SPL6/9/16 may be great significance to the construction of longan globular embryos.

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备注/Memo

备注/Memo:
收稿日期 Received: 2019-07-14 接受日期 Accepted: 2019-08-13
*国家自然科学基金项目(31672127)、福建省高原学科建设经费(102/71201801101)和福建农林大学科技创新基金(CXZX2017187和CXZX2018078)资助
**通讯作者Correspondence author (E-mail: buliang84@163.com)
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更新日期/Last Update: 2019-09-04