IGEM:Missouri University of Science and Technology/2009/Notebook/Cyanobacteria Mediated Filtration of Coal Flue/2013/09/27

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Date: 09/27/13 People in lab: Emily Puleo

Title: PCR hmp and norCB for TOPO TA cloning

Start Time: 10:15 AM

Purpose: To add prefix/suffix to hmp and norCB

Protocol: LTM ed. 2 Exceptions: Annealing Temp: 65C, Elongation time 7min

Products:

Sample Label Description
9/27/13 A norCB norCB with prefix/suffix
9/27/13 A hmp hmp with prefix/suffix
9/27/13 hmp F primer 1:100 dilution of hmp F primer
9/27/13 hmp R primer 1:100 dilution of hmp R primer

Notes: See 8/6/13 for hmp calculations, 7/31/13 for norCB. Restarted at 12:35PM

Stop Time: 3:15 PM

Next: TOPO TA cloning and transformation

Date: 09/27/13 People in lab: Emily Puleo, Levi Palmer

Title: TOPO TA cloning of hmp and norCB, gel electrophoresis to check for products of PCR

Start Time: 4:45 PM

Purpose: To check for prescence of genes and amplify by TOPO-TA cloning

Protocol: LTM ed. 2, Invitrogen TOPO-TA cloning manual

Products:

Sample Label Description
9/27/13 CT1 norCB transformed, 20uL plated
9/27/13 CT2 norCB transformed, 200uL plated
9/27/13 CT1 hmp transformed, 20uL plated
9/27/13 CT2 hmp transfored, 200uL plated

Results: hmp visible on gel, norCB had no product visible

Next: Check for hmp growth, miniprep if present. Redo PCR for norCB

Date: 09/27/13 People in lab: Emily Puleo, Levi Palmer

Title: Gel extraction and ligation of backbone

Start Time: 6:50 PM

Purpose: To extract the hmp from the gel, to convert the linear plasmid backbone to a circular backbone.

Protocol: LTM ed. 2

Products:

Sample Label Description
9/27 hmp GE hmp with iGEM prefix/suffix

Notes: Did not digest backbone first, cannot ligate.

Stop Time: 8:00 PM

Next: Digestion of backbone


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