1 / 10

LABORATORY 4A: Making Sure You’ve Got A Recombinant Plasmid

LABORATORY 4A: Making Sure You’ve Got A Recombinant Plasmid. 2014. Plasmid of interest, pARA-R. Restriction digest of pARA-R. Review of Lab 2A. R-tube contains pARA-R plasmid and no restriction enzymes (control) R+ tube contains pARA-R plasmid and restriction enzymes HindIII and BamH I

sharne
Download Presentation

LABORATORY 4A: Making Sure You’ve Got A Recombinant Plasmid

An Image/Link below is provided (as is) to download presentation Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author. Content is provided to you AS IS for your information and personal use only. Download presentation by click this link. While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server. During download, if you can't get a presentation, the file might be deleted by the publisher.

E N D

Presentation Transcript

  1. LABORATORY 4A: Making Sure You’ve Got A Recombinant Plasmid 2014

  2. Plasmid of interest, pARA-R

  3. Restriction digest of pARA-R

  4. Review of Lab 2A • R-tube contains pARA-R plasmid and no restriction enzymes (control) • R+ tube contains pARA-R plasmid and restriction enzymes HindIII and BamH I • Digested by incubating at 37°C for 60 minutes

  5. Goals of Lab 4A • Determine if restriction enzymes digested the pARA-R plasmids • Use gel electrophoresis to separate the digest products according to “size” • Allow visualization of restriction fragments and non-digested plasmids on the gel

  6. Running the gel Wells

  7. What to expect in the visualization • Restriction fragments are linear and their speed through the gel is only determined by size • Undigested plasmids have multiple configurations and their speed through the gel is determined by size and shape

  8. Plasmid configurations Supercoiled Nicked circle Multimer

  9. Predict the visualization

  10. Results DL R– R+ Multimer Nicked Supercoiled 5 Kb Linear Fragment Linear Fragment

More Related