Haynes Lab:Notebook/Engineering PC-TFs/2014/07/09: Difference between revisions

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----
----
'''Restriction Digest'''
'''Restriction Digest'''
{| {{table}} border="1" cellspacing="3"  
{| {{table}} border="1" cellspacing="5"  
|- valign="top"
|- valign="top"
| bgcolor=#cfcfcf | Reagent  
| bgcolor=#cfcfcf | Reagent  
| bgcolor=#cfcfcf | Volume
| bgcolor=#cfcfcf | Volume
| rowspan="7" | <u>Expected:</u><br>BL01(W1:2-4), BL05(W1:5, W2:2-3) <br>Forward insertion: 710, 7010. <br>Reverse insertion: 3230, 4490. <br>No insertion: 710, 4490.<br> BL09 (W2:4-6) <br>Forward insertion: 710, 5921 <br>Reverse insertion: 2141, 4490 <br>No insertion: 710, 4490.<br> *Final well is empty CMV/MV9.
| rowspan="9" | <u>Expected:</u><br>BL01(W1:2-4), BL05(W1:5, W2:2-3) <br>Forward insertion: 710, 7010. <br>Reverse insertion: 3230, 4490. <br>No insertion: 710, 4490.<br> BL09 (W2:4-6) <br>Forward insertion: 710, 5921 <br>Reverse insertion: 2141, 4490 <br>No insertion: 710, 4490.<br> *Final well is empty CMV/MV9.
| rowspan="7" | [[Image:W120140709 174255.jpg|400px|Today's gel]]<br>15 μL/lane; 1% agarose;<br> <u>Observed lengths:</u> W2: 7100, 700; W3: 7100, 5200, 2500; W4: *5200, 2500, 710; W5: 5000, 2500; W6: 5000, 2500, 710.<br> *Uncut plasmid.<br> [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
| rowspan="10" | [[Image:W120140709 174255.jpg|400px|Today's gel]]<br>15 μL/lane; 1% agarose;<br> <u>Observed lengths:</u> W2: 7100, 700; W3: 7100, 5200, 2500; W4: *5200, 2500, 710; W5: 5000, 2500; W6: 5000, 2500, 710.<br> *Uncut plasmid.<br> [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
| rowspan="7" | [[Image:W220140709 174229.jpg|400px|Today's gel]]<br>15 μL/lane; 1% agarose;<br> <u>Observed lengths:</u> W2: *7100, 700; W3: *6100, 2500, 710; W4: *7200, 2500, 710; W5: 5000, 4000; W6: *5500, 2500, 750.<br> *Uncut plasmid.<br> [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
| rowspan="10" | [[Image:W220140709 174229.jpg|400px|Today's gel]]<br>15 μL/lane; 1% agarose;<br> <u>Observed lengths:</u> W2: *7100, 700; W3: *6100, 2500, 710; W4: *7200, 2500, 710; W5: 5000, 4000; W6: *5500, 2500, 750.<br> *Uncut plasmid.<br> [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
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| DNA(BL01x3, BL05x3, BL09x3, CMV/MV9) || 5.0 μL
| DNA(BL01x3, BL05x3, BL09x3, CMV/MV9) || 5.0 μL
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'''Results'''
'''Results'''
<br>
<br>
Top W2: BL01 possible success.
Top W2: BL01 possible success.<br>
Bottom W2: BL05 possible success.
Bottom W2: BL05 possible success.<br>
Bottom W4: BL09 possible success.
Bottom W4: BL09 possible success.<br>
 
----
'''Sequencing Results Order 9218'''
<br>
<u>BL01rA<br></u>
*Used 1μL BL01 miniprep 1, 1μL DD123 B primer, 8μLdH<sub>2</sub>O.
*'''Results''': Matches were found after the primer and then just before CMV suggesting no insert.
<br>
<u>BL01rB</u><br>
*Used 1μL BL01 miniprep 2, 1μL DD123 B primer, 8μLdH<sub>2</sub>O.
*'''Results''': Matches were found after the primer and then just before CMV suggesting no insert.
<br>
<u>BL09rA</u><br>
*Used 1μL BL09 miniprep 1, 1μL DD123 B primer, 8μLdH<sub>2</sub>O.
*'''Results''': Matches were found after the primer and then just before CMV suggesting no insert.
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__NOTOC__
__NOTOC__

Latest revision as of 00:05, 27 September 2017



 		Main project page
Previous entry      Next entry

Summary

Liquid Culture Minipreps
Eluted with 70μL dH2O.

Plasmid OD260 OD260/280 ng/μL
1. BL01 C1 0.289 1.89 288.704
1. BL01 C2 0.463 1.918 462.857
1. BL01 C3 0.495 1.915 495.398


Plasmid OD260 OD260/280 ng/μL
1. BL05 C1 0.047 2.124 46.727
1. BL05 C2 0.301 1.912 301.14
1. BL05 C3 0.075 1.97 74.969


Plasmid OD260 OD260/280 ng/μL
1. BL09 C1 0.059 1.986 59.185
1. BL09 C2 0.236 1.878 235.782
1. BL09 C3 0.185 1.958 184.993


Restriction Digest

Reagent Volume Expected:
BL01(W1:2-4), BL05(W1:5, W2:2-3)
Forward insertion: 710, 7010.
Reverse insertion: 3230, 4490.
No insertion: 710, 4490.
BL09 (W2:4-6)
Forward insertion: 710, 5921
Reverse insertion: 2141, 4490
No insertion: 710, 4490.
*Final well is empty CMV/MV9. 		Today's gel
15 μL/lane; 1% agarose;
Observed lengths: W2: 7100, 700; W3: 7100, 5200, 2500; W4: *5200, 2500, 710; W5: 5000, 2500; W6: 5000, 2500, 710.
*Uncut plasmid.
Ladder 		Today's gel
15 μL/lane; 1% agarose;
Observed lengths: W2: *7100, 700; W3: *6100, 2500, 710; W4: *7200, 2500, 710; W5: 5000, 4000; W6: *5500, 2500, 750.
*Uncut plasmid.
Ladder
DNA(BL01x3, BL05x3, BL09x3, CMV/MV9) 5.0 μL
10X buffer 1.5 μL
XbaI 1.0 μL
NruI 1.0 μL
dH2O 6.5 μL
Total 15 μL --> 37°C/ 30 min.


Results
Top W2: BL01 possible success.
Bottom W2: BL05 possible success.
Bottom W4: BL09 possible success.

Sequencing Results Order 9218
BL01rA

  • Used 1μL BL01 miniprep 1, 1μL DD123 B primer, 8μLdH2O.
  • Results: Matches were found after the primer and then just before CMV suggesting no insert.


BL01rB

  • Used 1μL BL01 miniprep 2, 1μL DD123 B primer, 8μLdH2O.
  • Results: Matches were found after the primer and then just before CMV suggesting no insert.


BL09rA

  • Used 1μL BL09 miniprep 1, 1μL DD123 B primer, 8μLdH2O.
  • Results: Matches were found after the primer and then just before CMV suggesting no insert.
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