Difference: ProtocolsConjugation (11 vs. 12)

Revision 122020-03-02 - SeanLeonard

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META TOPICPARENT name="ProtocolList"

General Conjugation Protocol

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  1. Incubate conjugation plate overnight (See image 2 for example of overnight growth).
  2. Scrape up each conjugation mixture into a micro centrifuge tube with 1 mL PBS. Vortex and gently spin down. Repeat.
  3. Resuspend conjugation mixture in 1 mL of PBS. Plate 100 ÁL of this mixture and 100 ÁL of a 10-fold dilution onto selective plates.
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  1. Pick single colonies and restreak or grown in selective media. Confirm transfer of the plasmid via PCR amplification of the insert. conj_plates-01.png
    conj_plates-02.png
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  1. Pick single colonies and restreak or grown in selective media. Confirm transfer of the plasmid via PCR amplification of the insert.
conj_plates-01.png
conj_plates-02.png
 

Considerations and customization:

Many aspects of this protocol can be tweaked if your initial conjugation attempts are unsuccessful.
 
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