3208 pEPD-FW

Cloning of

Purpose:  

Date: 2018-6-22

Notebook: 

Page:

Clone ID:3208

Experimenters: Yijia, Paul 

Results

PCR

Date: Date: 6/20/2018

By: Yijia

template:3152 pCEH0-Flag-VH-mIgG2a  ;3010 pCMV5-Flag-kappa as template

primers:

Fragment 1: PCR from 3152 pCEH0-Flag-VH-mIgG2a as template

3208-F1 ACTCGCGGCCCAGCCGGCCATGgctgtacactccGAGGTTCAGCTGCAGCAGTC

3208-R1 aGATCCGCCACCGCCGGATCCACCTCCGCCTGAACCGCCTCCACCGCTAGCAGAGGACACGGTCAGTGTG (CACACTGACCGTGTCCTCTGCTAGCGGTGGAGGCGGTTCAGGCGGAGGTGGATCCGGCGGTGGCGGATCt)

Fragment 2: PCR from 3010 pCMV5-Flag-kappa as template

3208-F2 GATCCGGCGGTGGCGGATCt aga tct GCGCGC TGTGACGTGCTGATGACACAGGCTC

3208-R2 atggtgttggcctcccgggccTAGGACCtcgAGcttGgtAcctccgccaaaggTG

Gel:

1st PCR of 3208

2nd PCR of 3208

Cloning

By: Paul

Vector: 3200 pEPD digested with SfiI

Ligation: 30 min with T4 DNA ligase

Competent cells: NEBTurbo

Colony PCR Screening 

pADL-F and 3208-R2

Restriction Digestion

Date: 6/21/2018

By: Paul

Nco1 + Xho1

Sequencing

6/22/18, clone #1, no good, missing 1C

pick more clones, make moiniprep.

send #8 to sequencing

sequencing: clone #8 is good