Experiments This Week
Monday – Nanodropped the protein and found its concentration. Ran it on two 12% SDS-PAGE, stained and destained. The protein did not run in either of them.
Tuesday – Yoshie poured the gels, to make sure it wasn’t that. We ran BSA, cell lysate, before IPTG, rtcB. rtcB still did not run.
Wednesday – Nanodropped oligos and made stock solutions; ran 20% SDS-PAGE oligo gel; stained with SYBR gold and saw faint bands w/ UV. Attempted 5-fold dilutions of protein that did not affect anything
Thursday – ran two oligo SDS-PAGE gels: 20% and 15% precast; imaged on UV – much better bands – ran the oligos off the 15% gel; tried to make 10x TBE and failed
Friday – attempted imaging of previous 20% SDS-PAGE gel on typhoon (dye degraded); setup of account; planning of solubility experiments for rtcB
UV transilluminator – only absorbs – doesn’t need filter
Purchases for the week = $39.54 (rack, 4-way rack, cryo vials, 2 plastic media 1 L bottles)
What I Still Need to Do
1. Optimize buffer for protein solubility.
3. Find the article: Determination of monolayer-protected gold nanoparticle ligand-shell morphology using NMR. Liu, Xiang Yu, Miao; Kim, Hyewon; Mameli, Marta; Stellacci, Francesco Nat Commun; Volume: 3, Date: 2012 , Pages: 1182 2012
Tentative Schedule for Next Week
Monday – prepare two 12% SDS-PAGE gels, prepare 12 different buffers for rtcB, run them, stain and destain; prep 20% gel and oligo dilutions; run them, stain them, image on typhoon. Make 10x TBE.
Tuesday – run the protein gels; make more protein samples to try
Wednesday – redo the 20% oligo gel and redo typhoon
Thursday – (possible) express protein
Friday – (possible) purify protein