中文版 | English
题名

单细胞水平多染色质蛋白分析技术开发及应用

其他题名
DEVELOPMENT AND APPLICATION OF MULTI-CHROMATIN PROTEIN ANALYSIS TECHNOLOGY AT SINGLE CELL LEVEL
姓名
姓名拼音
CHEN Yiling
学号
12032144
学位类型
硕士
学位专业
0710 生物学
学科门类/专业学位类别
07 理学
导师
方亮
导师单位
前沿与交叉科学研究院
外机构导师单位
华大基因
论文答辩日期
2023-05-15
论文提交日期
2023-06-29
学位授予单位
南方科技大学
学位授予地点
深圳
摘要
  表观遗传研究技术可应用于组蛋白修饰及转录因子的研究,为剖析细胞特征和发育轨迹提供强有力的依据。但同时分析并比较多个染色质相关蛋白的特征仍然存在挑战性。本研究探索开发了一种可用于同时分析多个染色质相关蛋白表观基因组特征的新方法,既可以克服捕获结果的稀疏性问题,又可以获得细胞特异性表观基因组模式的重要信息。
  本研究基于 CUT&Tag 方法,设计制备了装载有组蛋白特异性标签的蛋白 A-Tn5 转座酶(pA-Tn5),将其与多种不同的抗体结合形成复合物;与细胞孵育后,每个抗体识别并结合到染色质上不同的组蛋白,随后 pA-Tn5 在目标组蛋白附近切割 DNA,并插入带有特异性标签的接头序列;回收标签化的 DNA 片段后,使用引物识别添加标签的序列进行 PCR 扩增,建库测序。这一方法使同时定位多个染色质相关蛋白成为可能。在鉴定染色质蛋白共定位时,本技术利用多种抗体-蛋白 A-Tn5 转座酶复合物所携带的特异性标签,将不同标签的接头序列插入在两个共定位的组蛋白附近区域的DNA 中,因此打断所得片段两端将包含不同抗体的标签,标记为混合 reads。通过混合 reads 与单个组蛋白定位区域的特异性 reads 的对比,实现了识别同一染色质区域的不同蛋白的共定位。通过单细胞液滴微流控系统,本研究还实现单细胞水平下多个不同组蛋白修饰区的信息捕获。
  综上所述,本研究开发的技术实现了在多细胞与单细胞水平下同时定位多种组蛋白的全基因组结合位点,该技术可以作为表观基因组研究的新工具,更加便捷地获取更丰富的组蛋白修饰信息。
关键词
语种
中文
培养类别
独立培养
入学年份
2020
学位授予年份
2023-06
参考文献列表

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所在学位评定分委会
生物学
国内图书分类号
Q81
来源库
人工提交
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条目标识符http://sustech.caswiz.com/handle/2SGJ60CL/544501
专题南方科技大学
生命科学学院_生物系
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陈依玲. 单细胞水平多染色质蛋白分析技术开发及应用[D]. 深圳. 南方科技大学,2023.
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