1439 pFus B10-SMN2-L2

Cloning of 1439 pFus B10-SMN2-L2

Purpose: Cloning of 1439 pFus B10-SMN2-L2

Date: 2012-1---

Notebook:0

Page:

Clone ID:1439

Experimenters:  shanyi

Results

Day2, pick white clones to PCR using primers pCR8_F1 and pCR8_R1 :

Day3

 Miniprep the  plasmids: Use enzymes AflII and XbaI to digest

Top1-5,Top1-8,Bottom1-1, Bottom2-1,Bottom2-4,Bottom2-5 may be right.

Transform TOP2 again.

Day4, pick white clones to PCR using primers pCR8_F1 and pCR8_R1 :

Day5

Mini prep the plasmids: Use enzymes AflII and XbaI to digest

Top2-8,Top1-12,Top1-14 may be right.

Sequence TOP1-5,BOTTOM1-1,BOTTOM2-1 and top2-8,top2-14 with primers pCR8_F1 and pCR8_R1.

TOP1-5,BOTTOM1-1,BOTTOM2-1 are right.

Top2-8,Top2-14 are empty vectors, so need to repeat step1.

TOP2 .
Repeat reaction1,digest and ligate again,

pick white clones to PCR using primers pCR8_F1 and pCR8_R1 :

TOP2 CHANGE F4 pFUS-B-10, C1,A6,D2,D4, do reaction#1 again. pick white clones to PCR using primers pCR8_F1 and pCR8_R1 : still no band(dna polymerase is not good)

Mini prep the plasmids: Use enzymes AflII and XbaI to digest

Sequence TOP2N-1 with primers pCR8_F1 and pCR8_R1.

Top2n-1 is right.