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特异性针对大鼠甲状旁腺激素受体1基因siRNA表达载体构建及鉴定 被引量:4

Construction and identification of siRNA recombinant expression vector targeting parathyroid hormone 1 receptor gene
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摘要 目的:构建并筛选携带针对大鼠甲状旁腺激素受体1基因的pSUPERretro-GFP/Neo干扰逆转录病毒载体及鉴定。方法:根据Genbank筛选甲状旁腺激素受体1基因3个阳性克隆及1个阴性克隆序列,定向克隆入逆转录病毒载体pSUPERretro-GFP/Neo,并用RT-PCR和测序鉴定,将其转染至胰岛素-1细胞中,Westernblot观察甲状旁腺激素受体1基因表达,鉴定其转染效率。结果:菌落RT-PCR及基因测序鉴定结果表明序列正确,并能成功转染到胰岛素-1细胞株中,Western blot筛选最佳抑制基因。结论:成功构建并筛选出特异性沉默大鼠甲状旁腺激素受体1基因的pSUPERretro-GFP/Neo干扰逆转录病毒载体,为深入探讨大鼠甲状旁腺激素受体1基因的抗凋亡作用奠定了试验基础。 Objective To construct and screen the expression vector of siRNA specific for parathyroid hormone 1 receptor gene.Methods Genome sequences of parathyroid hormone 1 receptor gene was retrieved from gene bank,and 4 pairs of oligonucleotides were synthesized and inserted into pSUPER.retro RNAi,which was identified by RT-PCR and sequence analysis,and then was transfected into INS-1 cells.The expression of parathyroid hormone 1 receptor gene was observed with Western blot and the transfection efficiency was identified.Results It was confirmed by RT-PCR and sequence analysis that siRNA recombinant expression vector targeting parathyroid hormone 1 receptor gene was coincided with the designs completely,then transfected into INS-1 cell successfully.The best gene was picked up by Western blot.Conclusion The successful construction of recombinant parathyroid hormone 1 receptor-siRNA vectors establishes a favorable foundation for further study on anti-apoptosis of the parathyroid hormone 1 receptor gene.
出处 《中华实用诊断与治疗杂志》 2010年第5期444-447,共4页 Journal of Chinese Practical Diagnosis and Therapy
基金 广西科技厅自然科学基金资助项目(991294) 广西北海市科学研究与技术开发计划项目(200901059)
关键词 甲状旁腺激素受体1基因 小分子干扰RNA 小发卡结构RNA 转染 RNA干扰 Parathyroid hormone 1 receptor gene small interfering RNA small hairpin RNA transfer RNA interference
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参考文献8

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共引文献4

同被引文献44

  • 1黄慧敏,张红,李惠翔,姜国忠,高冬玲,王秀利.表达E2F1小干扰RNA的逆转录病毒载体的构建[J].实用诊断与治疗杂志,2005,19(3):177-179. 被引量:5
  • 2刘小友,于立新,付绍杰,孙煦勇,袁谦,熊海云.大鼠肾移植急性排斥反应模型的建立[J].中华泌尿外科杂志,2005,26(11):730-732. 被引量:11
  • 3王康,戴勇,李德萍,王宇罡,吕天羽.大鼠原位肾移植模型肾静脉吻合方法的改良[J].中华器官移植杂志,2006,27(1):54-56. 被引量:11
  • 4张怀军,李桑蕾,张磊,黄磊,邢新,李蠡.病理性瘢痕组织中E2F1基因的表达及其意义[J].实用诊断与治疗杂志,2007,21(3):187-189. 被引量:8
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