Experimental Methods for Use with Lambda
Experimental Methods for Use with Lambda
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与 Lambda 一起使用的实验方法
DOI：
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发表时间：
1983
期刊：
影响因子：
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通讯作者：
K. Murray
中科院分区：
文献类型：
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作者：
W. Arber;L. Enquist;B. Hohn;N. Murray;K. Murray
引言本文特别针对（但不限于）那些使用λ作为载体克隆其他生物基因的人。在这篇文章中，我们描述了生长和纯化噬菌体的技术，并提供了一个简短的测试菜单，用于验证在一些广泛使用的载体中发现的遗传特征。我们还提出了简单的菌株构建，诱变，DNA纯化和体外DNA包装的方法。也可以找到替代和重叠的程序（米勒，1972;戴维斯等，1980;马尼亚提斯等，1982;伯曼等，1983）。我们强调，途径的选择往往是个人偏好的问题，方法可以而且应该变化，以适应特定的噬菌体菌株，实验室条件和方便的调查。我们已经指出了为什么某些操作是首选或避免的，但我们警告读者，原因往往是推测性的，并提出了去神秘化我们的主题，而不是抑制实验。细菌宿主菌株的生长和繁殖λ噬菌体生长的常用宿主是大肠杆菌K12的衍生物。表1列出了本文特别提到的几种菌株。E. coliK 12及其符号由Bachmann和Low（1980）列出。对于λ的滴定和繁殖，细菌最好在37°C通风的丰富培养基中生长，如胰蛋白胨肉汤，当它们达到一定的浓度时使用。
INTRODUCTION This paper is addressed particularly (but not exclusively) to those who use λ as a vector for cloning genes of other organisms. In it we describe techniques for growing and purifying phage and offer a brief menu of tests that are useful for verification of genetic characters found in some widely used vectors. We also suggest methods for simple strain construction, mutagenesis, DNA purification, and in vitro DNA packaging. Alternative and overlapping procedures can also be found (Miller 1972; Davis et al. 1980; Maniatis et al. 1982; Berman et al. 1983). We emphasize that the choice of route is often a matter of individual preference and that methods can and should be varied to suit the particular phage strain, laboratory conditions, and the convenience of the investigator. We have indicated why certain manipulations are to be preferred or avoided, but we warn the reader that the reasons are often speculative and are presented to demystify our subject rather than to inhibit experimentation. GROWTH AND STORAGE OF BACTERIAL HOST STRAINS The commonly used hosts for the growth of phage λ are derivatives of Escherichia coli K12. Those few strains to which particular reference is made in this paper are given in Table 1. All known genes of E. coli K12 and their symbols are listed by Bachmann and Low (1980). For the titration and propagation of λ , bacteria are best grown in a rich medium, such as tryptone broth, with aeration at 37°C, and used when they have reached a...