The direction and rate of bacteriophage T7 DNA packaging in vitro.
The direction and rate of bacteriophage T7 DNA packaging in vitro.
复制标题
噬菌体T7 DNA体外包装的方向和速率。
DOI：
10.1006/viro.1993.1476
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发表时间：
1993
期刊：
影响因子：
3.7
通讯作者：
Serwer,P
中科院分区：
文献类型：
--
作者：
Son,M;Watson,RH;Serwer,P
为了确定T7噬菌体DNA体外包装过程中DNA进入的方向，将不完全包装的DNA（ipDNA）降解到衣壳外，然后将包装的DNA从衣壳中释放出来，通过琼脂糖凝胶电泳进行分级。在分级分离后，用右端特异性寡核苷酸进行定量凝胶内探测检测异质ipDNA（称为右端ipDNA）。大多数右端ipDNA以成熟T7 DNA前体的预期动力学出现。使用左端特异性寡核苷酸的凝胶内探测检测ipDNA（左端ipDNA）;左端ipDNA的摩尔量总是比右端ipDNA的摩尔量少至少50倍。左侧末端ipDNA随着T7 DNA包装的流产终产物的动力学而出现。因此，生产性T7 DNA包装以从右到左的方向发生。对右端ipDNA转化为成熟长度DNA的定量产生了对从右到左的体外T7 DNA包装的平均速率的估计：对于包装的最后20-50%的DNA，为28 ± 6 kbp/min。
To determine the direction of the entry of DNA duringin vitrobacteriophage T7 DNA packaging, incompletely packaged DNA (ipDNA) was fractionated by agarose gel electrophoresis after degradation of DNA outside of capsids and then release of packaged DNA from capsids. After fractionation, quantitative in-gel probing with a right end-specific oligonucleotide detects heterogeneous ipDNA (called right-end ipDNA). Most of the right-end ipDNA appears with kinetics expected of a precursor to the mature T7 DNA. In-gel probing with a left-end-specific oligonucleotide detects ipDNA (left-end ipDNA); the molar amount of left end ipDNA is always at least 50× less than the molar amount of right-end ipDNA. Left-end ipDNA appears with the kinetics of an abortive end product of T7 DNA packaging. Thus, productive T7 DNA packaging occurs in a right-to-left direction. Quantitation of the conversion of right-end ipDNA to mature-length DNA yields an estimate of the mean rate of right-to-leftin vitroT7 DNA packaging: 28 ± 6 kbp/min for the last 20-50% of the DNA packaged.