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鸡肝脏染色质的表征及其体外转录产物的分析。

Characterization of chick liver chromatin and analysis of its in vitro transcription products.

作者信息

Dierks-Ventling C, Stalder J, Gautschi J

出版信息

Nucleic Acids Res. 1978 Jul;5(7):2643-56. doi: 10.1093/nar/5.7.2643.

DOI:10.1093/nar/5.7.2643
PMID:566911
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC342192/
Abstract

Carefully controlled preparation of chromatin from purified chick liver nuclei yielded over 50% native chromatin as shown by the analysis of the nucleosome pattern after micrococcal nuclease digestion. The size of DNA in this chromatin as analyzed on alkaline sucrose gradients varied from 10S to 19S, the majority being 14S. All endogenous RNA polymerases were represented in the chromatin preparation although to different extents: RNA polymerase I was the most and RNA polymerase II the least abundant. Initiation studies showed that endogenous RNA polymerase II was capable of initiating RNA chains during 5 min. Saturation of chromatin with purified homologous RNA polymerase II increased initiation to 10 min. The addition of heparin caused the RNA transcribed to be larger in size and of increased yield. Chromatin transcription with added purified RNA polymerase II in the presence of heparin produced RNA as large as 32S. A chromatin preparation of this kind would therefore be suitable to transcribe any eukaryotic gene invitro provided additional homologous RNA polymerase II is used.Images

摘要

通过精心控制从纯化的鸡肝细胞核制备染色质，如经微球菌核酸酶消化后对核小体模式的分析所示，可得到超过50%的天然染色质。在碱性蔗糖梯度上分析，这种染色质中DNA的大小从10S到19S不等，大多数为14S。尽管程度不同，但所有内源性RNA聚合酶都存在于染色体制备物中：RNA聚合酶I含量最高，RNA聚合酶II含量最低。起始研究表明，内源性RNA聚合酶II能够在5分钟内起始RNA链。用纯化的同源RNA聚合酶II使染色质饱和可将起始时间延长至10分钟。加入肝素会使转录的RNA尺寸更大且产量增加。在肝素存在下用添加的纯化RNA聚合酶II进行染色质转录可产生高达32S的RNA。因此，只要使用额外的同源RNA聚合酶II，这种染色体制备物就适合在体外转录任何真核基因。图片

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab41/342192/896fef1d2a97/nar00468-0424-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab41/342192/896fef1d2a97/nar00468-0424-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ab41/342192/896fef1d2a97/nar00468-0424-a.jpg

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Characterization of chick liver chromatin and analysis of its in vitro transcription products.鸡肝脏染色质的表征及其体外转录产物的分析。
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引用本文的文献

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Nucleic Acids Res. 1978 Oct;5(10):3929-43. doi: 10.1093/nar/5.10.3929.
2
Properties of in vitro transcription by isolated Xenopus oocyte nucleoli.非洲爪蟾卵母细胞核仁体外转录的特性
Nucleic Acids Res. 1979;6(5):1929-40. doi: 10.1093/nar/6.5.1929.

本文引用的文献

1
Preparation of native chromatin and damage caused by shearing.制备天然染色质和剪切引起的损伤。
Science. 1975 Mar 28;187(4182):1203-6. doi: 10.1126/science.187.4182.1203.
2
SEDIMENTATION STUDIES OF THE SIZE AND SHAPE OF DNA.DNA大小与形状的沉降研究
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The precision of ultraviolet absorption measurements in the Schmidt-Thannhauser procedure for nucleic acid estimation.施密特 - 坦豪泽核酸估测法中紫外线吸收测量的精度。
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Animal DNA-dependent RNA polymerases. 1. Large-scale solubilization and separation of A and B calf-thymus RNA-polymerase activities.动物依赖DNA的RNA聚合酶。1. 小牛胸腺RNA聚合酶A和B活性的大规模溶解与分离。
Eur J Biochem. 1972 Jul 13;28(2):269-76. doi: 10.1111/j.1432-1033.1972.tb01910.x.
7
Transcription of Xenopus chromatin by homologous ribonucleic acid polymerase: aberrant synthesis of ribosomal and 5S ribonucleic acid.非洲爪蟾染色质由同源核糖核酸聚合酶进行转录：核糖体核糖核酸和5S核糖核酸的异常合成
Biochemistry. 1974 Apr 23;13(9):1896-9. doi: 10.1021/bi00706a018.
8
Preparation of chromatin. Variation in the template properties of chromatin dependent on the method of perparation.染色质的制备。染色质模板特性的变化取决于制备方法。
Eur J Biochem. 1974 Aug 1;46(3):461-71. doi: 10.1111/j.1432-1033.1974.tb03639.x.
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Chromatin structure: a repeating unit of histones and DNA.染色质结构：组蛋白与DNA的重复单元。
Science. 1974 May 24;184(4139):868-71. doi: 10.1126/science.184.4139.868.
10
Spectrophotometric determination of protein concentration in cell extracts containing tRNA's and rRNA's.分光光度法测定含有转运核糖核酸和核糖体核糖核酸的细胞提取物中的蛋白质浓度。
Anal Biochem. 1973 Aug;54(2):454-63. doi: 10.1016/0003-2697(73)90374-6.