IGEM:Peking University/2008/Notebook/Group 1/2008/07/28

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PKU_080728_G1_1
Picking up colonies-lacS/tet

Enzyme cutting and gel running

Miniprep Biobrick

Report: Testing result-Xili's plasmid-pADH-lac1/2/S/tet
Digestion-pADH-lac1/2-1/2-2/S/tet-ClaI-BamHI

Gel running and testing

Gel Picture: 080725-xili_plasmid_test-redigest

lanes:1kb marker, 100bp marker, lac1, lac2-1, lac2-2, lacS, tet, lambda marker

Transformation-xili-lac1-1/1-2/2-1/2-2/S-1/S-2/tet-1/tet-2(from two envelops)

Picking up colonies|Miniprep|digestion(40min)|Gel running(140V)

Gel Picture: 080728-xili_plasmid_test-redigest

Lanes: lambda marker, {lac2-2-1, lac2-2-2, lac2-2-3, lac2-2-4, lac2-2-5, lac2-2-6, lacS-2-1}0727miniprep, {lac1-1-1, lac2-2-1, lac2-2-2, lac2-2-3, lacS-1-1, lacS-2-1}0726miniprep, 100bp marker

Picking up colonies|Miniprep|digestion(90min)|Gel running(120V)

Gel Picture: 080728-xili_plasmid_test-redigest-longer_time

Lanes: {lac2-2-1, lac2-2-2, lac2-2-3, lac2-2-4, lac2-2-5, lac2-2-6, lacS-2-1}0727miniprep, lambda marker, {lac1-1-1, lac2-2-1, lac2-2-2, lac2-2-3, lacS-1-1, lacS-2-1}0726miniprep

Conlusion:

1. The plasmids are right from the enzyme cutting result: only 700bp bands, no 1.4kb bands, showing the exact fragment we need.

2. The middle one looks nicer--nearly no smearing phenomena, which suggests the following aspects MAY be reasons: a, shorter enzyme cutting time (since BamHI has potential spark activity); b, higher gel running volt (time to run gel becomes shorter); c, new K buffer (other materials are totally the same, including the enzyme containers).

3. Longer time for digestion may generate some new bands, if not contamination in lane last but two of pic3...


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