Single nucleotide primer extension: quantitative range, variability, and multiplex analysis.

Single nucleotide primer extension: quantitative range, variability, and multiplex analysis.
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单核苷酸引物延伸:定量范围、变异性和多重分析。

DOI:
10.1101/gr.6.4.336
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发表时间:
1996
期刊:
影响因子:
7
通讯作者:
Burke,DT
Burke,DT
中科院分区:
生物学1区
文献类型:
--
作者:
Greenwood,AD;Burke,DT

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二倍体基因座同源等位基因转录产物的定量测定在哺乳动物基因表达研究中具有广泛的应用前景。单核苷酸引物延伸(SNuPE)分析是一种简便、灵敏的等位基因转录本鉴别方法,只需要等位基因之间1bp的差异。本研究对poly(dT)-引物和基因特异性逆转录的有效范围、实验变化和影响进行了研究。从单个逆转录反应中分析多个基因的能力也被评估。对于被检测的基因,当次要转录物占主要等位基因的1/250时,达到最大检测范围。测定内或测定之间的误差相对较小,响应的线性保持在大约千倍的范围内。
The quantitative measurement of transcription products from homologous alleles at a diploid locus has broad application for the study of mammalian gene expression. Single nucleotide primer extension (SNuPE) analysis is a simple and sensitive method for allelic transcript discrimination requiring only 1 bp difference between alleles. In this study the effective range, experimental variation, and the influences of poly(dT)-primed and gene-specific reverse transcriptions are characterized. The ability to analyze several genes from a single reverse transcription reaction is assessed as well. For the genes examined, the maximum range of detection is reached when the minor transcript represents 1/250 of the major allele. Relatively little error is seen within or between assays and linearity of response is maintained over an approximately thousandfold range.
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