1255 pTXB1-C6H2

Cloning of 1255 pTXB1-C6H2

Purpose: Cloning of 1255 pTXB1-C6H2

Date: July 6, 2012

Notebook: 

Page:

Clone ID: 1255

Experimenters: Jennifer Truong

Results

Cloning

Amplified from 1126 (pTXB1) template with pTXB1-Xho1-F and pTXB1-Pst-R2 primers

Cut out band in lane 4, expected size 624 bp, purified and digested  with restriction enzymes Xho1 and Pst1

Digested DNA and vector with restriction enzymes Xho1 and Pst1

Cut out bands from lane 3 and 4, purify, ligate and transform, plate and let grow overnight

Is lane 4 vector pTXB1? it should have two bands, one vector, one original insert.

Dropped DNA band, and vector band is a little too light, reamplified

Cut out lane 1 (using Phusion from Thermo) and 2 (using homemade Phusion), expected size 624 bp, purified and digested  with restriction enzymes Xho1 and Pst1,

Lane 1 and 2 are the digested DNA and vector with Xho1+ Pst1 (from Phusion from Thermo)

Lane 3 and 4 are the digested DNA and vector with Xho1 and Pst1 (from Homemade Phusion)

Lane 4 digestion does not look good (vector) 

Only cut out lane 1 and 2, purified and ligated together overnight. Paul transformed with Top10 cells and plated

Colony Screening

Date: June 9, 2012

Digested MiniPrep plasmids with restriction enzymes Xho1 and Pst1

May have postive insert for all samples, but unclear