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大鼠白蛋白结构基因序列的构建与克隆

Construction and cloning of rat albumin structural gene sequences.

作者信息

Kioussis D, Hamilton R, Hanson R W, Tilghman S M, Taylor J M

出版信息

Proc Natl Acad Sci U S A. 1979 Sep;76(9):4370-4. doi: 10.1073/pnas.76.9.4370.

DOI:10.1073/pnas.76.9.4370
PMID:291970
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC411576/
Abstract

A recombinant plasmid containing a DNA segment complementary to rat liver albumin mRNA has been constructed, cloned, and used to examine the organization of albumin gene. The 18S fraction of total liver poly(A)-containing RNA was copied into a double-stranded cDNA by avian myeloblastosis virus reverse transcriptase and Escherichia coli DNA polymerase I. The cDNA was inserted into the HindIII site of the plasmid pBR322 via the addition of specific oligonucleotide linkers. Recombinant plasmids were screened by hybrid arrest of mRNA translation and hybridization with specific cDNAs. Thereby, a plasmid was identified that contained a 1200-nucleotide insert corresponding to a segment adjacent to the 5'-terminal region of albumin mRNA. The inserted sequence was used as a hybridization probe to detect five EcoRI fragments of genomic DNA which encode albumin mRNA. These were compared to eight EcoRI fragments identified within the rat genome by albumin cDNA. We conclude that the albumin gene (or genes) is interrupted at more than one site in the coding DNA by intervening sequences. Furthermore, we were able to distinguish those fragments that encode the 5' and 3' ends of the mRNA.

摘要

构建了一个含有与大鼠肝脏白蛋白mRNA互补的DNA片段的重组质粒，进行了克隆，并用于研究白蛋白基因的组织。用禽成髓细胞瘤病毒逆转录酶和大肠杆菌DNA聚合酶I将肝脏总含poly(A)RNA的18S部分转录成双链cDNA。通过添加特定的寡核苷酸接头，将cDNA插入质粒pBR322的HindIII位点。通过mRNA翻译的杂交阻断和与特定cDNA的杂交筛选重组质粒。由此鉴定出一个质粒，其含有一个1200个核苷酸的插入片段，对应于白蛋白mRNA 5'-末端区域相邻的一段序列。将插入序列用作杂交探针来检测编码白蛋白mRNA的基因组DNA的五个EcoRI片段。将这些片段与通过白蛋白cDNA在大鼠基因组中鉴定出的八个EcoRI片段进行比较。我们得出结论，白蛋白基因（或多个基因）在编码DNA中的多个位点被间隔序列中断。此外，我们能够区分那些编码mRNA 5'和3'末端的片段。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/2888d02d1904/pnas00009-0221-c.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/12b772dcd3bb/pnas00009-0220-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/da4ff491dadb/pnas00009-0220-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/507b722edb67/pnas00009-0221-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/7fc337b47ea3/pnas00009-0221-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/2888d02d1904/pnas00009-0221-c.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/12b772dcd3bb/pnas00009-0220-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/da4ff491dadb/pnas00009-0220-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/507b722edb67/pnas00009-0221-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/7fc337b47ea3/pnas00009-0221-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/0cc9/411576/2888d02d1904/pnas00009-0221-c.jpg

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SEDIMENTATION STUDIES OF THE SIZE AND SHAPE OF DNA.DNA大小与形状的沉降研究
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A seven-base-pair deletion in an intron of the albumin gene of analbuminemic rats.无白蛋白血症大鼠白蛋白基因内含子中的一个七碱基对缺失。
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