DNA片段3'末端核苷酸的测定。
Determination of the 3' terminal nucleotide of DNA fragments.
作者信息
Olson K, Harvey C
出版信息
Nucleic Acids Res. 1975 Mar;2(3):319-25. doi: 10.1093/nar/2.3.319.
Abstract
A new method for determining the 3'-terminal nucleotide of a DNA strand is presented. Use is made of the fact that one (and only one) 2',3'-dideoxyribonucleotide can be added to the 3'-end of a DNA fragment with calf thymus terminal transferase. Addition of more than one nucleotide analog per strand is impossible due to the absence of a 3'-terminal hydroxyl group. If the terminatind dideoxyribonucleotide contains an (alpha 32p) label, the resulting 3'-blocked strand can be digested by "nearest neighbor" techniques and the original 3'-endgroup determined. Picomole quantities of DNA strands can be labeled and the 3'-end determined.
摘要
提出了一种确定DNA链3'-末端核苷酸的新方法。利用了这样一个事实：用小牛胸腺末端转移酶可以将一种（且仅一种）2',3'-双脱氧核糖核苷酸添加到DNA片段的3'-末端。由于不存在3'-末端羟基，每条链添加多个核苷酸类似物是不可能的。如果终止双脱氧核糖核苷酸含有（α-32P）标记，则所得的3'-封闭链可以通过“相邻核苷酸”技术进行消化，并确定原始的3'-末端基团。可以对皮摩尔量的DNA链进行标记并确定3'-末端。
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