發(fā)布時間：2018-06-01 08:13

  本文選題：核轉(zhuǎn)錄因子紅細胞系相關(guān)因子- + RNA干擾�。� 參考：《臨床神經(jīng)病學(xué)雜志》2017年02期

【摘要】：目的設(shè)計及構(gòu)建小鼠核轉(zhuǎn)錄因子紅細胞系相關(guān)因子-2(Nrf2)基因的干擾質(zhì)粒,并篩選出效果最好的干擾質(zhì)粒。方法設(shè)計3組針對Nrf2基因的核糖核酸干擾(RNAi)序列,應(yīng)用基因重組技術(shù)克隆入載體中構(gòu)建短發(fā)夾RNA(shRNA),分別為shRNA1、shRNA2及shRNA3,通過基因測序鑒定,經(jīng)Lipofectamine2000轉(zhuǎn)染至BV2細胞,實時PCR檢測Nrf2 mRNA的表達,Western Blot法檢測Nrf2蛋白的表達。結(jié)果測序表明,克隆入載體中的Nrf2干擾序列及讀碼框完全正確,實時PCR及Western Blot顯示shRNA3干擾效果最強。結(jié)論成功構(gòu)建小鼠Nrf2的有效干擾質(zhì)粒,為Nrf2信號通路在腦卒中領(lǐng)域的功能研究奠定了基礎(chǔ)。
[Abstract]:Objective to design and construct the interference plasmid of mouse nuclear transcription factor RBC related factor -2nrf2, and to screen out the best interference plasmid. Methods three sets of RNA interference RNAi (RNAi) sequences targeting Nrf2 gene were designed and cloned into the vector by gene recombination technique to construct shRNA2 and shRNA3 for short hairpin RNAs, which were identified by gene sequencing and transfected into BV2 cells by Lipofectamine2000. The expression of Nrf2 mRNA was detected by real time PCR and the expression of Nrf2 protein was detected by Western Blot. Results sequencing showed that the Nrf2 interference sequence and the reading frame cloned into the vector were completely correct, and real-time PCR and Western Blot showed the best effect on shRNA3 interference. Conclusion the effective interference plasmid of mouse Nrf2 was successfully constructed, which laid a foundation for the study of the function of Nrf2 signaling pathway in the field of stroke.
【作者單位】： 江蘇大學(xué)附屬宜興醫(yī)院(揚州大學(xué)醫(yī)學(xué)院宜興臨床學(xué)院)神經(jīng)內(nèi)科;
【基金】：江蘇省科技廳自然科學(xué)基金面上項目(BK20141121) 無錫市衛(wèi)生局青年基金(Q201301;Q201625)
【分類號】：R743.3
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本文編號：1963493