Map of Experiments for Phage Library

2015-01-13

azim58 - Map of Experiments for Phage Library

gel extraction 6-2-09
PCANTAB5E 6-5-09
Transformation with X2 cut PCANTAB5E 6-8-09
RNA Isolation 6-9-09
Competent Cells 6-15-09
Transformation to check competent cells 6-17-09
phage display preparation 6-19-09
phage display 6-19-09
amplified eluate 6-23-09
rna extraction 6-30-09
transformation with scFv plasmid (primer calculations) 7-6-09
Spleen Extraction 7-8-09
RNA extraction 7-8-09
RNA gel 7-8-09
cDNA synthesis 7-9-09
cDNA gel 7-10-09
RNA extraction 7-11-09
synthetic scFv Library 7-12-09
synthetic library rescue 7-15-09 to 7-16-09
separating phage from cells 7-18-09
phage titer 7-18-09
phage titer results 7-19-09
phage titer calculations 7-20-09
helper plasmid (M13cp) Transformation 7-23-09
preparing electrically competent cells 7-24-09
m13 plasmid transformation 7-25-09
competent cells 7-26-09 to 7-27-09
m13 transformation with chloramphenicol 7-28-09
ER2738 Transformation 7-29-09
transformation of dh10b with pLACI 7-29-09
Lower concentration chloramphenicol 7-30-09
analyzing competent cell results 7-31-09
scFv plasmid transformation 7-31-09
transformation of helper plasmids 8-1-09
electrical competent cell preparation 8-2-09
Midi of M13cp 8-3-09
Electroporation of several plasmids 8/4/09
biotinylate ab 8/5/09
phage ELISA 8-5-09 to 8-6-09
Phage ELISA Results 8-7-09
Dot Blot Analysis 8-7-09
Dot Blot Results 8-7-09
Biotinylation of ab after removal of sodium azide 8-12-09
dot blot to check biotinylation 8-13-09
dot blot w/hrp 8-13-09
dot blot imaging 8-15-09
dot blot procedure plan 8-13-09
results of m13cp gel 8-18-09
phage ELISA 8-19-09 to 8-20-09
m13 plasmid glycerol stock 8-21-09
phage ELISA Plan 8-21-09
Dilution series of alexafluor 8-21-09
Phage ELISA Results 8-24-09 to 8-25-09
phage ELISA Results 8-25-09
biotinylation of phage ab 8-28-09
Phage ELISA 9-1-09
Phage ELISA Results 9-1-09
Estimating Phage Concentration from absorbance 9-3-09
m13cp verification 9-3-09
titration of scFv 9 9-3-09
Results of scFv9 Titration 9-6-09
Phage on Array schematic 9-7-09
phage on array calculations 9-7-09
phage array protocol 9-17-09
naive library plasmid prep 9-17-09
preparing to make electrically competent m13cp 9-22-09
phage mice immunization 9-23-09
preparation of electrocompetent e coli protocol 9-24-09
preparation of electrocompetent m13cp and m13cp-CT 9-24-09
transformation of naive library into helper strains 9-25-09
retransformed library into helper strains 9-26-09
check on naive lib into helper cell transformation 9-28-09
glycerol stock to phage protocol 9-29-09
glycerol stock of phage 9-29-09
glycerol stock to phage 10-1-09
titration of 6 monoclonal phages 10-5-09
result of 6 monoclonal phage titration 10-6-09
2nd phage mice immunization 10-7-09
m13cp verification 2nd attempt 10-7-09
plasmid extraction w/spin minikit 10-8-09
plasmid extraction w/qiagen kit 10-9-09
sequencing monoclonal phage 10-9-09
restriction digest of helper plasmids 10-9-09
planning naive library amplification 10-12-09
naive library amplification plan 10-13-09
mouse cheekbleed/naive library amplification 10-14-09
monoclonal phage sequence analysis/naive library amplification

10-15-09

naive library titration result 10-17-09
naive library titration 10-19-09
anti-M13 Mouse sera ELISA w/HRP 10-19-09
anti-M13 Mouse sera results 10-19-09
cancer B cell isolation 10-21-09
mouse sera phage ELISA preparation 10-21-09
mouse sera phage ELISA 10-21-09
mouse sera phage ELISA results 10-21-09
3rd phage mouse immunization 10-23-09
naive library verification 10-22-09
cancer b cell rna extraction 10-22-09
helper and PCANTAB gel 10-26-09
anti-m13 mouse sera ELISA 10-29-09
mouse sera ELISA results 10-30-09
pcr planning 11-2-09
anti-m13 mouse euthanization 11-4-09
cr of m13cp-CT-2 sample 11-6-09
result of m13cp-CT-2 PCR 11-9-09
pcr of m13cp-1 and m13cp-2 11-9-09
gel extraction of m13cp-1 11-12-09
phage insert check pcr results 11-13-09
ELISA for mouse sera titer 11-13-09
ELISA Mouse sera titer results 11-13-09
rna extraction schematic 3-8-10

Tumor and B cell RNA Extraction 6-14-10
RNA Isolation from B cells of Naive Mouse 6-14-10
Tumor, spleen, and blood collection 6-18-10
Isolation of B cells 6-18-10
Isolation of RNA 6-21-10
Naive B RNA Check 6-22-10
RNA Isolation from Tumor Tissue 6-24-10
Rerun of tumor RNA Gel 6-25-10
Preparation of cDNA from B cell RNA 7-1-10
Beta actin check of cDNA 7-2-10
Amplification of Antibody Gene Regions from cDNA 7-6-10 to 7-7-10
PCR of Antibody Gene Region 7-8-10
cDNA Synthesis 7-8-10
Naive Spleen Processing 7-19-10
RNA from Naive B cells 7-19-10
cDNA from Naive B RNA 7-20-10
PCR to Amplify antibody gene regions 7-21-10 to 7-22-10
Gel of Antibody Genes 7-23-10
Rerun of PCR to amplify ab genes 7-26-10 to 7-26-10
test PCR of ab genes 7-29-10
gel check of ab gene PCR 7-30-10
RNA Stability Test 7-30-10
PCR of Tumor ab genes 8-2-10
test of pcr with Hojoon's cDNA 8-3-10
pcr of ab genes with lower primer conc. 8-4-10
pcr of ab genes with more template 8-5-10
process spleens of smc1 mice 8-9-10
optimization of pcr of ab genes 8-10-10
optimization of pcr of ab genes 8-10-10
amplifying with all 68 primers for naive 8-10-10
pcr ab genes (tumor, smc1, 6-21) 8-17-10 to 8-21-10
pcr ab genes (tumor, smc1, 6-21) with extra primer and polymerase to

amplify faint bands

pcr ab genes gel images 8-21-11
extra thermocycles for rest of pcr plates 8-23-10
summary of pcrs 8-23-10
cDNA synthesis of smc1 RNA 8-24-10
amplification of smc1 ab genes 8-25-10
additional pcr cycles for troublesome primer pairs 8-27-10
pcr to amplify genes with difficult primers 8-27-10
gel extraction of ab genes 8-30-10
pcr purification of ab genes 8-30-10
pcr purification of ab genes 8-31-10
conclusion of analysis of ab genes 9-8-10
addition of linker to 1-78 9-9-10
addition of linker to 1-78 9-9-10
gel extraction of ab genes w/linker 9-10-10
overlapping pcr 9-10-10
flanking primers for overlapping pcr 9-11-10
check overlapping pcr and amplification of tumor ab genes 9-12-10
pcr to add linker to 1-78 genes 9-13-10
gel extraction of ab genes with linker 9-14-10
DH10B competent cell prep 9-15-10
Amplification of linked heavy and light 9-15-10
check of linked heavy and light 1-78 ab genes 9-16-10
check of tumor ab genes 9-16-10
pcr purification of tumor ab genes 9-20-10
check of transformation efficiency 9-20-10
gel extraction of tumor ab genes 9-21-10
naive ab gene amplification and first 10 tumor ab gene amplification

9-21-10

additional thermocycles for naive and tumor genes 9-22-10
gel of naive ab genes and 1st 10 tumor ab genes 9-23-10
tumor ab genes 7-10 9-23-10
DH10B competent cell preparation 9-24-10
gel extraction and pcr purification 9-29-10
nanodrop concentrations of naive 9-29-10
pcr for last few tumor ab genes 10-1-10
pcr purification and gel extraction 10-2-10
pool of primers w/SMC1 DNA 10-4-10
pcr of gel extracted ab smc1 gene 10-6-10

Construction of tumor scFv DNA 10-7-10 to 10-8-10
Amplification of Tumor scFV DNA 10-10-10
Construction of naive, SMC1 and 1-78 scFV DNA 10-11-10
Link heavy and light chains of 1-78 Naive and SMC1 DNA 10-13-10
Amplification of linked heavy and light chain genes 10-14-10
Amplification of scFV DNA 10-15-10
Restriction of 1-78 scFV with sfiI 10-18-10
Better picture of scFV DNA 10-20-10
Final scFV DNA gel 10-22-10
Final scFV DNA gel with fresh buffer 10-25-10
Gel extraction of antibody genes with linker 10-27-10
Construction of scFV 10-29-10
Gel extraction of final scFV 10-30-10
Amplified linked scFV fragments 11-2-10
scFV construction 11-4-10
Tumor lambda construction 11-5-10 to 11-8-10
Gel of final scFV DNA 11-11-10
Restriction digest of PCANTAB5E 11-15-10 to 11-16-10
Isolation of scFv DNA 11-18-10
Investigation of PCANTAB5E 11-17-10

Ligate PCANTAB with rCompTT 4-8-11
Ethanol precipitate ligation 4-11-11
Preparation of phage helper competent cells 4-19-11 to 4-21-11
Transformation of helper competent cells 4-22-11
Preparation to make competent cells 4-26-11
Preparation of Electrocompetent ER2738 w/M13cp-CT 4-29-11
Transformation efficiency and glycerol stock 5-4-11
Colony PCR for F1 ori 5-24-11
Helper ER2738 W/M13cp-CT preparation 7-20-11
Transformation of ER2738 M13cp-CT 7-26-11
Electrocompetent ER2738 M13cp-CT preparation 8-2-11
Test transformation "fast freeze H2O" 8-3-11
Electrocompetent cell preparation 8-4-11
Electrocomporation of ER2738 8-5-11
Transformation results 8-6-11
Test ligation for ER2738 8-11-11
EtOH precipitation and transformation 8-12-11
Cut PCANTAB5E 8-16-11
Recut PCANTAB for commercially competent cells 8-18-11
Gel extraction and ligation 8-19-11
Ethnal precipitation and transformation 8-20-11
Sfi I mouse SMC1fs 8-25-11
Restriction digest table 8-25-11
PCR purification/Gel extraction 8-25-11
Ethanol precipitation 8-26-11
Eletroporation of mouse SMC1fs scFV 8-26-11
Minipreps SMC1fs scFV 8-28-11
Colony PCR of SMC1fs scFV 8-29-11
Colony PCR test 9-1-11
Advantage PCR of miniprep 9-1-11
Cut SMC1fs scFV PCANTAB plasmids 9-6-11
BamHI and Hind III double digest 9-7-11
Restriction Digest and overnight Gel 9-7-11
Not I restriction digest check 9-10-11
Hind III & EcoRI digest 9-9-11
Not I digest with both buffers 9-9-11
Basic field inversion electrophoresis test 9-10-11
Cut PCANTAB for stuffer (SacI and PstI) 9-15-11
Cut PCANTAB with SacI and NotI 9-19-11
pGEX-SMC1fs cut with NotI 9-22-11
pRSET cut with SacI and NotI 9-24-11
Gel of pRSET sequential cut 9-24-11
Ligation 9-24-11
Ethanol precipitation PCANTAB pRSET 9-26-11
Electroporation protocol 9-26-11
tRNA- carrier preparation 10-3-11
PCANTAB -PRSET digest 10-4-11
Potential PCANTAB pRSET 10-5-11
Ethanol precipitation of PCANTAB maxiprep 10-19-11

Check of PCANTAB Maxiprep 11-1-11
PTAB Fragments into pJET 11-3-11
PCR of PCANTAB pJET clones 11-7-11
Gel and Miniprep 11-8-11

Cut Vector with SpeI and NotI 061412
Modification of pComb3xSS vector 6-25-12

o Diagram of nucleotide changes 6-26-12

Cut pComb with SfiI and NotI 7-6-12
PCR of pComb fragment 7-7-12
pComb In-Fusion 7-9-12
Resequence pComb NotI Modification 7-19-12
Miniprep pComb Cultures 7-21-12
added polyT near lacI in pComb 8-27-12
pcomb mod with 2nd round primers 8-29-12
modified pcomb electroporation 8-31-12
mpComb Sequencing 9-5-12
recut MN pComb 9-18-12 (MN is for modified NotI)
pComb fragment PCR round 1 9-19-12
pComb fragment PCR round 2 9-20-12
pComb In-Fusion 9-21-12
Extracted mpComb Plasmids 9-23-12
Cut pComb and rCompTT 10-4-12
NotI Digestion 10-5-12
ligate practice fragment 10-9-12
test for pComb Insert 10-15-12
pComb insert gel 10-15-12
pComb digest 10-16-12
CIAP treatment 10-16-12
pComb Gel extraction 10-16-12
pComb ligation 10-19-12
transformation of pcomb ligation 10-24-12
pcomb ligation and transformation 10-25-12
verify rcompTT ligation 10-29-12
rCompTT Ligation verification 10-29-12
sequence new pComb plasmid 10-31-12
pComb digest 10-16-12
CIAP Treatment 1-16-12
pComb Gel Extraction 10-16-12
pComb Ligation 10-19-12
Transformation of pComb Ligation 10-24-12
pComb Ligation and transformatio 10-25-12
verify rCompTT Ligation 10-29-12
rCompTT ligation verification 10-29-12
sequence new pcomb plasmid 10-31-12
pComb MN linearization 11-13-12
Construction of fragment for pComb 11-16-12
pComb MN linearization 11-16-12
Stellar electroporation 11-20-12
pComb linearization check 11-20-12
linearization with more polymerase 11-21-12
check of linearization 11-22-12
polymerase system test 11-26-12
check polymerase test 11-27-12
pComb linearization check 11-27-12
pComb linearization with PrimeSTAR 11-28-12
In-Fusion with linearized pComb 12-4-12
pComb relinearization 12-13-12
pComb linearization 1-14-13
In-fusion of linearized pComb 1-15-13
linearized pcomb gel extraction 1-16-13
sequencing pcomb candidates 1-18-13
sequenced pComb 1-29-13

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Note there seems to be some dates missing here such as "Cut rCompTT
w/SfiI 2-24-11)