Shared Laboratory Notebook

See also See http://2014.igem.org/Team:SF_Bay_Area_DIYbio/Notebook

For experiments from January 2016 on, see Experiments started January 2016

General Protocols

• DNA Handling
• Electra cloning protocol (Used for cloning IDT gBlocks into plasmid backbone)
• Media:Zymo E. coli competent cells transformation protocol.pdf (Protocol to transform plasmid DNA into chemically competent E. coli)
• Media:D4025i.pdf (Zyppy plasmid midiprep protocol; used to purify plasmid DNA from E. coli)
• Media:Clontech UTF-8'en-us'Quick Easy Yeast Transformation Mix Protocol-At-A-Glance 040214.pdf (Protocol to transform plasmid DNA into yeast cells)
• Sequencing Analysis using Serial Cloner 2-6-1
• Media:5-FOA Protocol, Zymo Research.pdf (Using 5-FOA to select against URA3+ yeast strains)
• Media:LifeTech_NuPAGE_TrisGlycine_Mini_Gels_Cat-EC6025BOX.pdf‎ (General protocol for precast Tris-Glycine 4%-20% acrylamide protein gels)

General Documentation Information

• Real Vegan Cheese Notebook Documentation Guidelines
• Real Vegan Cheese Notebook Training Sign-off

June 2014

• CR_062214_humanKappaCasein_001

August 2014

• CLONE_082514_humKappaCasein(Kex+)
• Growth Curve Experiment

September 2014

• Cloning 7 casein gBlocks, 12Sep2014 and on
• Troubleshooting failed 12Sep2014 transformation of P.FAKS.humBeta.S
• Cloning bovine beta casein, Fam20C (Kex + & -), from 27Sep2014
• Submitting 10 casein and FAM20C in pD1214 plasmids for sequencing 30Sep2014

October 2014

• Initial studies of yeast strains' growth conditions and troubleshooting
• Repeat Midipreps to select new clones for sequencing for failed sequences from Oct. 2
• Cloning human alpha casein S1, 04Oct2014 and on
• Checking for contamination in yeast strains, media, water, and lab surfaces 06Oct2014
• Cloning bovineKappa and bovineBeta into pSB1C3 Biobrick using NEB biobrick kit 08Oct2014
• Submitting 4 repeat and 1 new casein in pD1214 plasmid for sequencing 10Oct2014
• Cloning bovineKappa and bovineBeta into pSB1C3 Biobrick using NEB biobrick kit 10Oct2014
• Cloning bovineBeta into pSB1C3 Biobrick using NEB biobrick kit 13Oct2014
• Submitting FAM20C (Kex + & Kex -) for sequencing with new internal primers 14Oct2014
• Submitting FAM20C Kex + clone 9A and bovineKappa and bovineBeta: pSB1C3 Biobrick for sequencing 15Oct2014
• Electra cloning of #5 Casein insert 22Oct2014
• Experiment to determine whether yeast is contaminated and to salvage strain using 5FOA, 23Oct2014
• Test yeast transformation Bovine alpha S1 and Bovine beta B caseins 26Oct2014

November 2014

• Streaked new colonies of Bovine alpha S1 and Bovine beta B casein yeast transformations 03Nov2014
• Quantified IDT DNA (human kappa casein kex-) sample on nanodrop spectrophotometer to verify that the resuspension back in September worked. 06Nov2014
• Checked for colony growth in 03Nov2014 streak plates. Result: Yes 06Nov2014
• Making liquid cultures, in duplicate, of bovine Alpha S1 and bovine Beta B transformed yeast colonies. 12Nov2014
• Attempt SDS Page and also mini prep to confirm that yeast transformations worked 14Nov2014
• Repeat Mini Prep of bovine alpha S1 and bone beta B yeast 17Nov2014
• Yeast Transformation (Clontech kit) 18Nov2014
• SDS Page-Gel for all the bovine proteins 24Nov2014
• New 5 FOA Plates and prepare media 25Nov2014

December 2014

• Repeat SDS Page-Gel for all the bovine proteins 07Dec2014

January 2015

• Yeast Transformation of All Bovine Casein Inserts 10Jan2015
• SDS Page-Gel for all the bovine casein inserts 17Jan2015

May 2015

• Western Blot of bovine casein 2May2015

May 2016

• Plating URA- and Turbo Yeast Control 9-10 May 2016

June 2016