Plazmid DNA Qiagen Miniprep Kit kullanarak Bakteri kolonileri Arındırıcı

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

E. coli plazmid DNA arıtma moleküler klonlama için bir temel tekniktir. En az 5 ml bakteri kültürü Küçük ölçekli arıtma (miniprep) daha fazla enzimatik reaksiyonları (polimeraz zincir reaksiyonu ve restriksiyon enzimi sindirim) tarafından takip klon doğrulama ya da DNA izolasyonu, hızlı bir yoludur. Burada, moleküler biyoloji teknikleri için genel başlayanlar için bu son derece etkili bir tekniktir tanıtmak için amaçlayan QIAGEN QIAprep 8 Miniprep Kiti ile miniprep genel prosedürler, video kaydedildi. Tüm prosedürü yüksek tuz varlığı silika üzerine DNA adsorpsiyon takip E. coli hücrelerinin alkali lizis dayanır. 1) hazırlanması ve takas QIAprep membran, 3) yıkama ve plazmid DNA elüsyon üzerine DNA adsorpsiyon bakteriyel lizat, 2): Bu üç adımdan oluşur. Tüm adımlar, fenol, kloroform, CSCL, etidyum bromür, kullanmadan ve alkol yağış olmadan yapılmaktadır. Genellikle tüm prosedürü tamamlamak için en az 2 saat sürer.

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