Haynes Lab:Notebook/Engineering PC-TFs/2015/01/28

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Summary

Summary

Digest and Gel Verification

Reagent Volume

Expected:
BL01/5 Ligation Colonies [XbaI/NruI]
Forward insertion: 3659, 4064
Reverse insertion: 1538, 6185
No insertion: 1538, 4064

CMV/MV9 with [XbaI/NruI]
1538, 4064
Uncut CMV/MV9: 5197
Uncut CMV/BL05/MV9: 7723

Today's gel

15 μL/lane; 1% agarose;
Observed lengths:
W1: Ladder
W2: Ryan's stuff
W3: BL05 LCR Colony 11
W4: Colony 13
W5: Colony 17
W6: Colony 19
W7: BL05 Traditional Ligation Colony 32
W8: Colony 33
W9: Colony 36
W10: Colony 37
Ladder

DNA 8.0 μL
10X buffer 1.5 μL
XbaI 1.0 μL
BsmbI 0.5 μL
dH2O 4 μL
Total 15 μL --> 37°C/ 20 min.


Rather inconclusive without empty CMV/MV9. Again I'm not sure why several lanes appear empty when they have high concentration and purity given plate reader data. Will re-run with empty CMV/MV9 as wells 5&6 could have forward insertion (or uncut CMV/MV9 & no insertion bands).



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