Transformation of Plasmid DNA into E. coli Using the Heat Shock Method

Biology

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Froger, A., Hall, J. E. Transformation of Plasmid DNA into E. coli Using the Heat Shock Method. J. Vis. Exp. (6), e253, doi:10.3791/253 (2007).

Abstract

Transformation of plasmid DNA into E. coli using the heat shock method is a basic technique of molecular biology. It consists of inserting a foreign plasmid or ligation product into bacteria. This video protocol describes the traditional method of transformation using commercially available chemically competent bacteria from Genlantis. After a short incubation in ice, a mixture of chemically competent bacteria and DNA is placed at 42°C for 45 seconds (heat shock) and then placed back in ice. SOC media is added and the transformed cells are incubated at 37°C for 30 min with agitation. To be assured of isolating colonies irrespective of transformation efficiency, two quantities of transformed bacteria are plated. This traditional protocol can be used successfully to transform most commercially available competent bacteria. The turbocells from Genlantis can also be used in a novel 3-minute transformation protocol, described in the instruction manual.

Comments

14 Comments

  1. I have never seen or heard anyone use beads to create a spread plate. Is that a new technique that has not reached my university yet, or is it an outdated technique that scientists don't use anymore?

    Reply
    Posted by: Anonymous
    February 1, 2008 - 7:57 PM
  2. People use it all the time. Ask in the labs around, I am sure you will find people who use it.

    Reply
    Posted by: Anonymous
    February 1, 2008 - 8:11 PM
  3. I haven't seen it either so you're not alone.I'm wondering if it isn't easily to use a spreader in terms of cleanup and reducing contamination? 

    Reply
    Posted by: Anonymous
    March 9, 2008 - 8:45 PM
  4. Copacabana method for spreading E. coli and yeast colonies
    Mark T. Worthington, Roger Qi Luo, and Jared Pelo
    BioTechniques Vol. 30, No. 4: pp 738-741 (Apr ²001)

    Reply
    Posted by: ozgur g.
    September 9, 2008 - 9:04 PM
  5. chemical dna transformation and plasmid curing of E.coli

    Reply
    Posted by: Anonymous
    January 11, 2012 - 11:49 PM
  6. why don't the flame is open? Don't you consider the contamination risk, or do you think that it is not necessary??

    Reply
    Posted by: Anonymous
    September 9, 2008 - 4:22 PM
  7. Please look more carefully. The bunsen burner is clearly on and the blue flame is clearly visible in the video. It is on at all stages which are succeptible to contamination.   

    Reply
    Posted by: Anonymous
    September 16, 2008 - 8:57 PM
  8. That was indeed a great thing for the beginners in this field. Good job!

    Reply
    Posted by: Anonymous
    October 22, 2008 - 8:11 PM
  9. hi my neam isalirezababazade . iam student in unversity tehran . what do you teransfer factors F(negativ)in the e.coli?why transfe geneboth factor?

    Reply
    Posted by: Anonymous
    December 24, 2008 - 9:41 AM
  10. double stranded plasmid can be fntered into host

    Reply
    Posted by: Anonymous
    June 10, 2010 - 6:00 AM
  11. Beads are used as well as scoops, i prefare scoops because i can be sure that everything is spread nice and equal. some technician use beads, you need to do the right motion to make it work :D

    Reply
    Posted by: Anonymous
    January 25, 2012 - 3:59 AM
  12. Why we are keep the mixture of competent cells and plasmid DNA at 4²c for 45 minutes?

    Reply
    Posted by: Anonymous
    February 22, 2012 - 8:56 AM
  13. 45 seconds not minutes XD
    Useful, more protocols should be explain like this one
    thaks

    Reply
    Posted by: Anonymous
    April 12, 2012 - 7:48 PM
  14. Where is gloves to prevent contamination?????

    Reply
    Posted by: OHIONET O.
    May 10, 2012 - 4:35 PM

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