本文選題：釀酒酵母 + 細(xì)胞破碎　； 參考：《實驗室研究與探索》2016年08期

【摘要】：建立了一種簡便適用于從真核細(xì)胞釀酒酵母菌中提取總蛋白的方法,應(yīng)用該方法探索了兩株對核苷類藥物5-FC高耐受菌株的蛋白表達(dá)特性。實驗中以細(xì)胞破碎率和總蛋白濃度為主要檢測指標(biāo),對比分析了6種酵母細(xì)胞破碎方法,即液氮研磨、超聲波破碎、酸洗玻璃珠法、酶法、反復(fù)凍融法、TNBIO高壓細(xì)胞破碎,通過比色法測定了總蛋白的含量,發(fā)現(xiàn)改進(jìn)后的酸洗玻璃珠法的細(xì)胞破碎率為81.25%,總蛋白含量152.67 mg/m L,效果明顯優(yōu)于其他幾種方法.實驗表明,在裂解液中添加適量的PMSF等蛋白酶抑制劑,可以有效防止蛋白的降解。經(jīng)過SDSPAGE對核苷類藥物高耐藥性菌株酵母總蛋白表達(dá)特性分析,酸洗玻璃珠法提取的蛋白含量高,所含雜質(zhì)少,條帶比較多且清晰,其中相對分子質(zhì)量約為21 kDa和43kDa的蛋白有明顯的表達(dá)差異,可能與菌株細(xì)胞的耐藥性相關(guān)。
[Abstract]:A simple and convenient method for extracting total protein from eukaryotic Saccharomyces cerevisiae was established. The protein expression characteristics of two strains with high tolerance to nucleoside drugs 5-FC were investigated. In the experiment, the cell breakage rate and the total protein concentration were used as the main indexes to compare and analyze six kinds of yeast cell breaking methods, namely, liquid nitrogen grinding, ultrasonic crushing, acid washing glass bead method, enzymatic method, repeated freezing thawing method and TNBIO high pressure cell fragmentation. The content of total protein was determined by colorimetry. It was found that the cell fragmentation rate of the improved acid pickling glass bead method was 81.25 and the total protein content was 152.67 mg/m / L. the effect was obviously superior to that of other methods. The results showed that protease inhibitors such as PMSF could effectively prevent protein degradation. After SDSPAGE analysis of the expression characteristics of total protein in yeast with high resistance to nucleoside drugs, it was found that the protein extracted by acid washing glass bead method had a high content, less impurities and more bands. The protein with relative molecular weight of about 21 kDa and 43 kDa was significantly different, which may be related to the resistance of strain cells.
【作者單位】： 東華大學(xué)化學(xué)化工與生物工程學(xué)院;
【基金】：中央高�；究蒲袠I(yè)務(wù)費專項資金項目(2232014A3-03) 上海市自然科學(xué)基金項目(15ZR1400200)
【分類號】：R915

【相似文獻(xiàn)】

相關(guān)期刊論文 前8條

1 甄永蘇,彭澤,薛玉川;潘生丁降低核苷類藥物的毒性[J];生理科學(xué);1984年Z1期

2 瞿瑤;核苷類藥物抗肝炎病毒的態(tài)勢[J];中西醫(yī)結(jié)合肝病雜志;2002年04期

3 劉利;;治療慢性乙型肝炎的新型核苷類藥物[J];中國實用醫(yī)藥;2008年35期

4 潘贊紅;盧誠震;趙桂鳴;;抗HBV新核苷類藥物的臨床研究評價[J];天津藥學(xué);2007年05期

5 張振圖;郭富清;;核苷類藥物Epstein-Barr病毒(EBV)的研究[J];四川生理科學(xué)動態(tài);1986年02期

6 姚勤偉,黃春,吳昊;HIV-1逆轉(zhuǎn)錄酶耐核苷類藥物突變新進(jìn)展[J];國外醫(yī)學(xué).流行病學(xué)傳染病學(xué)分冊;2004年06期

7 邢利和;朱傳琳;;拉米夫定及其它核苷類藥物抗HBV的作用[J];傳染病信息;1999年02期

8 ;[J];;年期

相關(guān)重要報紙文章 前4條

1 遼寧錦州市傳染病醫(yī)院主任醫(yī)師 王振坤;核苷類藥物該吃多久[N];健康報;2007年

2 本報記者 賈巖;核苷類藥物邁入強(qiáng)效時代[N];醫(yī)藥經(jīng)濟(jì)報;2012年

3 北京地壇醫(yī)院 蔡fg東;核苷類藥物知多少[N];健康報;2007年

4 閆麗新;國產(chǎn)阿德福韋酯上市[N];健康報;2005年

，

本文編號：2084076