1513-pT7A3-polyAPol

Cloning of 1513-pT7A3-polyAPol

Purpose: Cloning of 11513-pT7A3-polyAPol

Date: 2013-2-25---2013-2-

Notebook:0

Page:

Clone ID:1513

Experimenters: Shanyi

Results

	band 1	band 3
	1279pT7A3	1140 pTXB1-polyAPol
enzymes	Nde1+ Xho1

Colony Screening

Date: 2013-2-

Make miniprep and digest with Nde1+ Xho1

chose 1513-2 to do protein expression screening ,cannot express.

subcloning

subclone polyAPol from 1140 pTXB1-polyAPol to 1279 pT7A3 using Nco1+ Xho1

http://52.1.104.161/eln/images/4/4c/Digest_for_1513.JPG

Subcloning and Screening

Digested by Nde1 and Xho1

Result intepretation/Summary

• <br>

Protein Expression

2013/3/11

Lane	Name	Protein Name	Clone ID	Host	Induction	Purification	pI/Mw
M	Invitrogen SeeBlue plus2
1	pT7A3-T4_RNA_Ligase_1	T4_RNA_Ligase_1	1049	5 ml, Nico21(DE3)plysN	0.2 mM IPTG  RT o/n	Ni beads	4.63 / 47485.21
2	pT7A3-MBP-(3C)-T4-RNA-Lig2	T4-RNA-Lig2	1123	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.11 / 39375.95
3	pT7A3-Klenow	Klenow	1501	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.11 / 71294.91
4	pT7A3-T4_RNA_Ligase_ 2_Truncated	T4_RNA_Ligase_ 2_Truncated	1503	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.14 / 29824.85
5	pT7A3-DNA-Pol-1	DNA-Pol-1	1508	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.40 / 103118.12
6	pT7A3-Phi29DNAPol	Phi29DNAPol	1512	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	8.61 / 66714.05
7	pT7A3-polyAPol	polyAPol	1513	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	9.67 / 53870.91
8	pT7A3-Klenow(3'->5'_exo–)	Klenow(3'->5'_exo–)	1537	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.77 / 67969.77
9	pT7A3-MBP-(3C)-Lambda-Exo	Lambda-Exo	1557	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.36 / 25925.62
10	pT7A3-MBP-(3c)-Endonuclease VIII	Endonuclease VIII	1572	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	7.77 / 29845.13

Protein Expression 2

2013/3/29
SHANYI

Lane	Name	Protein Name	Clone ID	Host	Induction	Purification	pI/Mw
M	PageRuler Plus Prestained protein ladder
1	pT7A3-T4-G32	T4-G32	1097	5 ml, Nico21(DE3)plysN	0.2 mM IPTG  RT o/n	Ni beads	4.82 / 33506.43
2	pT7A3-MBP-(3c)-MutM	MBP-(3c)-MutM	1138	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	8.68 / 30290.07+43.6
3	pT7A3-KOD	KOD	1458	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	7.53 / 90022.42
4	pT7A3-UDG	UDG	1494	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.45 / 27174.73
5	pT7A3-T7-Endo1	T7-Endo1	1505	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	9.54 / 17171.84
6	pT7A3-polyAPol	polyAPol	1513	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	9.67 / 53870.91
7	pT7A3-T7-Exo	T7-Exo	1558	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	4.93 / 34502.20
8	pT7A3-MBP-(3C)-Exo-1	MBP-(3C)-Exo-1	1562	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	8.80 / 18143.72
9	pT7A3-MBP-(3C)-RnaseH	MBP-(3C)-RnaseH	1573	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	8.43 / 17596.99 +43.6
10	pT7A3-MBP-(3c)-E_coli_Endo IV	MBP-(3c)-E_coli_Endo IV	1593	5 ml, Nico21(DE3)plysN	0.2mM IPTG RT o/n	Ni beads	5.43 / 31479.53 +43.6
11	pT7A3-T3 DNA ligase	T3 DNA ligase	1654	5 ml, Nico21(DE3)plysN	0.2 mM IPTG RT o/n	Ni beads	5.14 / 39350.70