1673-pET28b-NdeI

Cloning of 1673-pET28b-NdeI

Purpose: Cloning of 1673-pET28b-NdeI

Date: 2013-3-4---2013-3-7

Notebook:0

Page:

Clone ID:1673

Experimenters: yingying

Results

cloning

pcr1+pcr2

Colony Screening

Date: 2013-3-7

Make miniprep and digest with Nde1+Xho1

Result intepretation/Summary

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Conclusion

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Cloning of 1673-pET28b-NdeI

Purpose: Cloning of 1673-pET28b-NdeI

Date: 2013-3-11---2013-3-14

Notebook:0

Page:

Clone ID:1673

Experimenters: yingying

Results

cloning

fix the problem:
PCR using
Fragment 1
i_NdeI1-1+i_NdeI2-10

Fragment 2
i_NdeI2-11 + i_NdeI2-14

Gel purify, then join two fragments using

i_NdeI1-1 +i_NdeI2-14

Colony Screening

Date: 2013-3-14

Make miniprep and digest with Nde1+Xho1

protein screening

lane	name	clone ID	host	induction	purification		MW
1	pT7A3-T5-Exo	1567	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads		21KD/5.95
2	pCold-TF-(3c)	1614	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads	55.9KD/5.19
3	pET28b-Nde1	1673	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads		44KD/8.41
4	pTCold-MBP-(3C)	1677	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads		44KD
5	pCold-C(3C)-MBP-6H	1692	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads		46KD/5.77
6	pT7A3-wtSM2	1694	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads, should use CBD beads		33KD/6.30
7	pCold-MBP-(3C)-TEV	1696	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads		72KD/6.04
8	pCold-GST-(3c)-wt3C	1697	4 ml,NiCO21(DE3)pLysN	0.2 mM IPTG 25 oC o/n	Ni beads		21KD/8.47
9	pT7A3-MBP-(E3C)-p53wt	1707	4 ml, BL21(DE3)Star, pRare2-pLysS	0.2 mM IPTG 25 oC o/n	Ni beads		88KD/5.76
10	pET28b-hINFA2	1711	4 ml, BL21(DE3)Star, pRare2-pLysS	0.2 mM IPTG 25 oC o/n	Ni beads		21KD/5.95