Team:Macquarie Australia/WetLab/Notebook

Welcome to our Notebook! This page has been created to detail our projects progress since its conception. Click on the section titles below to expand pages that contain information regarding what we worked on and achieved each week. For a more detailed summary of what we achieved please visit the Results page.

Week 1

Biobrick stocktake of 2013 iGEM Macquarie_Australia parts: 11/11/13

• ChlG - sufficient plasmid stock
• DVR1 - sufficient plasmid stock
• ChlM - sufficient plasmid stock
• ChlI2 - need more plasmid stock
• POR- need more plasmid stock
• YCF - need more plasmid stock
• Plasto - need more plasmid stock
• GUN4- need more plasmid stock
• CTH1 - need more plasmid stock
• ChlD - need more plasmid stock. Question whether the 2013 part is really the reported sequence - something appears to be missing.

Send all for re-sequencing to verify DNA sequence as per registry entries.
DVR1 re-tranformation: Was re-done using gibson assembly and then transformed.

Week 2

Sequencing Results: all parts except ChlD were correct.

ChlD Fix: ChlD is missing 50 bp. Strategy to correct is to use ApaI and MluI restriction enzymes to cut out 50bp from clone of ChlD in pET vector from Willows group and re-insert into our BioBrick vector.

ApaI and MluI were used in a single digest according to manufacturer's instructions and as per ligation protocol on methods wiki. Fragments run on 1% agarose and gel purified. However, digestions were incomplete as viewed on agarose gel. Need to do separate digests for next attempt.

Increase stocks : Did plasmid preps to get more of: ChlI1; ChlI2; YCF54; ChlP, DVR1; POR

ChlH Biobrick correction

Attempt to make ChlH (BBa_K1080001) using combination of gblocks and PCR products, as designed by Macquarie_Australia 2013 iGEM team.

Assembly strategy is: G-Block –1 (470bp) + PCR-1 (304bp) + G-Block-2 (499bp) + G-Block-3 (499bp) + PCR-2 (984bp) + G-Block-4 (500bp) + G-Block-5 (481bp) + PCR-3 (673bp)

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