使用Qiagen公司的小量制备试剂盒的细菌菌落纯化质粒DNA

Biology

Your institution must subscribe to JoVE's Biology section to access this content.

Fill out the form below to receive a free trial or learn more about access:

Welcome!

Enter your email below to get your free 10 minute trial to JoVE!





We use/store this info to ensure you have proper access and that your account is secure. We may use this info to send you notifications about your account, your institutional access, and/or other related products. To learn more about our GDPR policies click here.

If you want more info regarding data storage, please contact gdpr@jove.com.

 

Summary

Cite this Article

Copy Citation | Download Citations

Zhang, S., Cahalan, M. D. Purifying Plasmid DNA from Bacterial Colonies Using the Qiagen Miniprep Kit. J. Vis. Exp. (6), e247, doi:10.3791/247 (2007).

Please note that all translations are automatically generated.

Click here for the english version. For other languages click here.

Abstract

大肠杆菌质粒DNA纯化是分子克隆的核心技术。小规模小于5毫升细菌培养纯化(小量)为进一步酶促反应(聚合酶链反应和限制性内切酶消化),克隆验证或DNA分离的快捷方式。在这里,我们录像通过QIAGEN的QIAprep 8小量制备试剂盒小量的一般程序,旨在引进这种高效的技术,分子生物学技术的一般初学者。整个过程是基于碱裂解大肠杆菌细胞,在高盐存在的DNA吸附到硅。它包括三个步骤:)1,编制和结算的一种细菌裂解液,2)的DNA吸附到QIAprep膜,3)洗涤和质粒DNA洗脱。所有步骤都没有使用苯酚,氯仿,氯化铯,溴化乙锭,无酒精沉淀。它通常需要不到2个小时,以完成整个过程。

Comments

6 Comments

  1. A very helpful and clear demonstration. Thanks!

    Reply
    Posted by: Anonymous
    January 22, 2008 - 9:33 PM
  2. With this kit is it best to do an alcohol precipitation after or unnecessary?   Thank you

    Reply
    Posted by: Anonymous
    November 4, 2008 - 7:26 AM
  3.   Hello from QIAGEN, There is no need to do an alcohol precipitation after this procedure - the DNA is ready to use as eluted from the QIAprep column. You can see the handbook for this kit online here if you would like:     http://www1.qiagen.com/literature/handbooks/literature.aspx?id=1000²48 -QIAGEN Technical Service     http://www1.qiagen.com/Contact/QIAGENWorldWide.aspx

    Reply
    Posted by: Anonymous
    November 4, 2008 - 8:18 AM
  4. By using this kit, what can I do if I'd like to increase the yield of plasmid with a low copy number? What is the maximum volume of culture that can be harvested?
    Thank you very much.

    Reply
    Posted by: Soon Keat O.
    July 18, 2009 - 10:51 AM
  5. Your demonstration is very good but you didn't mentioned the volumes of buffer added and the speed and time of centrifuge

    Reply
    Posted by: Anonymous
    March 2, 2010 - 10:40 AM
  6. Thanks for doing that video. There's not a lot of theory in there, but it is surely helpful for beginners. I'm holding onto it for future coworkers.

    Reply
    Posted by: Anonymous
    March 23, 2010 - 5:02 AM

Post a Question / Comment / Request

You must be signed in to post a comment. Please or create an account.

Usage Statistics