| 单炳辉，谢 鑫，李 季，柴普今，李鹤鹏，张琳贺，翟璐璐.蓝莓 Vco-miR_n10 的克隆及对干旱的响应分析[J].广东农业科学,2019,46(4):34-41 |
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蓝莓 Vco-miR_n10 的克隆及对干旱的响应分析 |
| Cloning of Blueberry Vco-miR_n10 and Analysis of Its Response to Drought |
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| DOI：10.16768/j.issn.1004-874X.2019.04.006 |
| 中文关键词: 蓝莓 拟南芥 miRNA Vco-miR_n10 甘露醇 |
| 英文关键词: blueberry Arabidopsis thaliana miRNA Vco-miR_n10 mannitol |
| 基金项目:国家自然科学基金（31701895） |
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| 摘要点击次数: 1834 |
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| 中文摘要: |
| 【目的】MicroRNA（miRNA）是一类内源单链非编码的小分子 RNA，在动植物中参与转录后基因
表达调控。Vco-miR_n10 是蓝莓中的新型 miRNA，目前尚未见该 miRNA 功能方面的报道。本研究对蓝莓 VcomiR_n10 进行了初步功能分析。【方法】基于前期蓝莓小 RNA 高通量测序结果，对鉴定到的蓝莓新型 miRNA
（Vco-miR_n10）的前体序列（224 nt）进行克隆与测序，并利用 mfold 在线软件对其前体序列的二级结构进行
预测。采用 Gateway 系统构建了 35Spro:: pre-Vco-miR_n10 表达载体并转入野生型拟南芥中，对其正常生长及 300
mmol/L 甘露醇处理条件下的表型进行观察与分析。【结果】克隆获得了 Vco-miR_n10 的前体序列，该序列能够折
叠形成较完美的茎环结构且仅有 2 个错配碱基对。Vco-miR_n10 的成熟体序列位于 3' 茎臂上。观察正常生长条
件下的转基因植株并无显著的表型变化。而在 300 mmol/L 甘露醇模拟的干旱条件下，Vco-miR_n10-24 与 VcomiR_n10-52 的发芽率（33.53%、37.69%）和绿苗率均显著低于野生型（发芽率 67.72%、绿苗率 66.33%）。【结
论】蓝莓 Vco-miR_n10 能够响应甘露醇模拟的干旱胁迫，并降低拟南芥对干旱的耐受能力，影响发芽率和绿苗率，
说明蓝莓 Vco-miR_n10 可能在植物响应干旱胁迫过程中发挥一定的调控功能。 |
| 英文摘要: |
| 【Objective】MicroRNAs (miRNAs) are a class of endogenous, single-stranded and non-coding small
RNA molecules that are involved in the regulation of gene expression after transcription in plants and animals. Vco-miR_
n10 is a novel miRNA in blueberry, and its function remains unknown. In the present study, a preliminary functional analysis
of blueberry Vco-miR_n10 was carried out.【Method】Based on the previous data from high throughput sequencing of
small RNA in blueberry, the precursor sequence (224 nt) of the novel miRNA (Vco-miR_n10) was cloned and sequenced.
The secondary structure of its precursor sequence was predicted by mfold web server. The expression vector, 35Spro::preVco-miR_n10, was constructed by Gateway system and transferred into wild type Arabidopsis thaliana and its phenotype
under the conditions of normal growth and 300 mmol/L mannitol treatment was observed and analyzed, respectively.【Result】
The precursor sequence of Vco-miR_n10 was obtained by cloning, and structure analysis indicated that the precursor shows
near-perfect stem-loop structure with no more than 2 mismatches. In addition, the mature sequence was located on the 3’stem arm. No significant phenotypic variation was observed in transgenic plants under the normal growth condition. However, under
the drought condition simulated by 300 mmol/L mannitol, the germination rate (33.53%, 37.69%) and cotyledon greening rate
(33.53%, 37.69%) of two transgenic lines (Vco-miR_n10-24 and Vco-miR_n10-52) were significantly lower than those of
wild type A. thalian (germination rate: 67.72%; cotyledon greening rate:66.33%).【Conclusion】These results indicated that
blueberry Vco-miR_n10 could respond to the mannitol-simulated drought stress, and reduce the drought tolerance in A. thalian
through affecting seed germination and cotyledon greening, which showed that blueberry Vco-miR_n10 might play a regulatory
role in plant response to drought stress. |
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