840:153g:Projects/project10

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Team Members

  • Lorena Christensen
  • Gabe Martin
  • Michelle Fretheim

Project Name and Description

  • Remediation of Phenol Resin using White Rot LiP transferred into E. Coli
  • Proposal:
     Group formed with interest of breaking down plastic as final goal.

1. Subsequent searches of the internet using //www.ncbi.nlm.nih.gov and Google scholar led to discovery of organism Phanerochaete chrysosporium, which presents phenol degrading capabilities through lignin peroxidase.

2. Obtain specimen white rot from Dr. Bumpus's lab. Extract DNA from mycelium using protocol by John J. Weiland.

3. Accession number for the gene of interest is EF644562.

4. First stage amplify gene using PCR, will need to build forward and reverse primers for the gene as well as for five introns needing to be removed. Cut with restriction enzymes and run on gel electrophoresis to check for removal of introns.

5. Build biobrick using format E X Part of Interest S P

6. Selection of vector for use, will need to use specific one for admission to biobricks registry

7. Use restriction enzymes to cut vector and gene of interest, ligate into plasmid

8. Place into E. Coli specimen, breed, observe traits in offspring

9. To verify successful inheritance will need to suppress levels of N, C, S in media in order to release repression of desired lignin peroxidase, substitute phenol as nutrient source. If trait is inherited will see discoloration in plastic in approximately two weeks. Will need to watch for possible death of offspring from process of breaking down phenol.

Final Presentation

Media:presentationreal.ppt

Media:Blastsequence.doc

Media:Sequences.txt

Important Results and Milestones

  • keep track of your most important results and refer to the corresponding page in your notebook
  • upload important pictures (don't forget to label them! Powerpoint is very convenient). Remember: these will become quite handy later in your summary report or final presentation. If you do label and upload the pictures as soon as you got them, your summary report can be written much more effortlessly (do you usually procrastinate? This is chance to do some work before hand that frees you up for finals week).

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