‘潘那利’番茄碱性螺旋环螺旋转录因子基因的克隆与表达分析
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中国博士后科学基金面上资助(2019M663862);


Cloning and Prokaryotic Expression Analysis of a Transcription Factor Gene in Solanum pennellii
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    摘要:

    碱性螺旋环螺旋(basic/HelixLoopHelix, bHLH)转录因子是植物最大的转录因子家族之一,其广泛参与植物逆境胁迫响应。该研究从野生‘潘那利’番茄(Solanum pennellii Correll)中成功克隆出bHLH转录因子基因SpbHLH89(Sol Genomics登录号Sopen04g001150),采用qRTPCR分析其在干旱胁迫下的表达模式,并利用异源表达初步分析其对非生物胁迫的响应。结果表明:(1)SpbHLH89编码区包含684 bp,编码227个氨基酸,具有典型的碱性螺旋环螺旋区,主要定位于细胞核中;进化树结果显示,SpbHLH89转录因子高度保守,与拟绒毛烟草NtbHLH94(Nicotiana tomentosiformis)存在高度相似性。(2)qRTPCR结果显示,SpbHLH89在‘潘那利’番茄的茎、叶和花中均有表达,其表达量受干旱胁迫诱导。(3)SDSPAGE与Western bloting结果显示,pET30aSpbHLH89重组蛋白大小约为31 kD。(4)在盐胁迫(400 mmol/L NaCl)和干旱胁迫(600 mmol/L甘露醇)条件下,异源表达重组蛋白的E. coli BL21(DE3)重组菌生长速度提高,说明异源表达SpbHLH89转录因子基因可提高细菌对非生物胁迫的耐受性。

    Abstract:

    Basic helixloophelix (bHLH) transcription factors are one of the largest transcription factor families in plants. It is extensively involved in stress response of plants. In the present study, we cloned a bHLH transcription factor geneSpbHLH89 (Sol Genomics number Sopen04g001150) from Solanum pennellii. Meanwhile, we also analyzed its expression patterns under drought stress by qRTPCR and verified its biological function by prokaryotic expression. The results showed that: (1) an open reading frame (ORF) of SpbHLH89 was obtained from S. pennellii, which is 684 bp long and encodes 227 amino acids with typical basichelixbasic domain and mainly located on the nucleus. Phylogenetic tree analysis showed that SpbHLH89 was highly conserved and shared a high degree of sequence similarity with NtbHLLH094 from Nicotiana tomentosiformis. (2) qRTPCR analysis showed that the pattern of SpbHLH89 was highly expressed in flowers and also induced by drought stress. (3) SDSPAGE and Western blotting analysis showed that the molecular weight of pET30aSpbHLH89 was approximately 31 kD. (4) Heteroexpression of the pET30aSpbHLH89 could improve the growth of the recombinant strain of E. coli BL21(DE3) under salt (400 mmol/L NaCl) and drought (600 mmol/L mannitol) stress. Taken together, the heterologous expression of SpbHLH89 transcription factor could improve the tolerance of recombinant bacteria to abiotic stress.

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胡佳蕙,王柏柯,李 宁,等.‘潘那利’番茄碱性螺旋环螺旋转录因子基因的克隆与表达分析[J].西北植物学报,2022,42(4):541-548

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  • 在线发布日期: 2022-05-18
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