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一种用于克隆稀有转录本5'末端的高效策略，可分离大鼠色氨酸羟化酶mRNA的多个5'非翻译区。

An efficient strategy for cloning 5' extremities of rare transcripts permits isolation of multiple 5'-untranslated regions of rat tryptophan hydroxylase mRNA.

作者信息

Delort J, Dumas J B, Darmon M C, Mallet J

机构信息

Laboratoire de Neurobiologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique, Gif-sur-Yvette, France.

出版信息

Nucleic Acids Res. 1989 Aug 25;17(16):6439-48. doi: 10.1093/nar/17.16.6439.

DOI:10.1093/nar/17.16.6439
PMID:2780283
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC318339/
Abstract

The 5' end mapping of rat tryptophan hydroxylase (TPH) mRNA indicated a diversity in 5'-untranslated regions. Corresponding sequences were isolated by a variant of the Polymerase Chain Reaction, recently designated as 'anchor PCR', and a 'cRNA enrichment' procedure. The latter circumvents the limitations of 'anchor PCR', which failed to yield minor TPH sequences: this novel strategy allows purification of specific DNA fragments by elimination of the unspecific products, generated by the PCR, which prevent further amplification. Analysis of TPH sequences strongly suggests that TPH mRNAs are synthesized from at least two promoters, the proximal one exhibiting two 'CCAAT homologies'.

摘要

大鼠色氨酸羟化酶（TPH）mRNA的5'端图谱显示其5'-非翻译区存在多样性。通过聚合酶链反应的一种变体（最近被命名为“锚定PCR”）和一种“cRNA富集”程序分离出了相应序列。后者克服了“锚定PCR”的局限性，“锚定PCR”无法产生少量的TPH序列：这种新策略通过消除PCR产生的非特异性产物来纯化特定的DNA片段，这些非特异性产物会阻止进一步扩增。对TPH序列的分析强烈表明，TPH mRNA至少由两个启动子合成，近端启动子表现出两个“CCAAT同源性”。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6d17/318339/bd04dc53a98f/nar00133-0021-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6d17/318339/2b907c87ad8d/nar00133-0019-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6d17/318339/bd04dc53a98f/nar00133-0021-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6d17/318339/2b907c87ad8d/nar00133-0019-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/6d17/318339/bd04dc53a98f/nar00133-0021-a.jpg

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