Team:Stony Brook/Notebook/Week6
Week 6
7/03:
• Nanodrop Results from plasmid miniprep
• Restriction Enzyme Digest of NEB Constructs
• Ran digests on 1.2% agarose gels, all failed except MBP
• PCR of Aureocin A 53, LnqZ, LnqZ-LnqQ hybrid, and Epidermicin NI01
• PCR Products run on 1.8% agarose gel
• PCR Purification of all constructs that ran properly on gel
7/04:
• Happy Independence Day!
• Nanodrop Results from PCR Purification
• Tried to PCR Aureocin A53 again, ran on agarose gel, failed
• Miniprep of MBP, GST, SUMO liquid cultures
• Glycerol stocks created of MBP, GST, SUMO cultures- stored in -80°C
• PCR Purification of Aureocin A53 R1
7/05:
• Nanodrop Results from PCR Purification: Aureocin A53: 88.5 ng/ul
• Nanodrop Results from miniprep
• Restriction Enzyme Digest of vectors and inserts
• Ran on a gel, everything was successful
• Gel Extracted Restriction Digest products
• Nanodrop Results of Gel Extracts
• PCR Purification Using Enzo and Qiagen Kits
• Transformed lnqZE and lnqZA tagged and untagged hybrids into DH5α E. coli
7/06:
• Miniprepped plasmid vectors
• Nanodrop Results from miniprep
• Gel Extract of LnqZQ hybrids, Nanodrop Results of Gel Extract
• Restriction Enzyme Digest of miniprepped vectors, messed up gel, so no results
• Ligation of inserts with GST and SUMO vectors (20 uL reactions)
• Set up reactions as shown below
• Gently pipette up and down and microfuge briefly
• Incubate at 16°C overnight
• PCR Purification Using Enzo Kit
• Made liquid cultures of lnqZE and lnqZA tagged and untagged hybrids
7/07:
• Transformed and plated single bacteriocin constructs in GST and SUMO vectors
• Retransformed lnqZE and lnqZA untagged hybrids
• Nanodrop Results from miniprep
