Haynes Lab:Notebook/David Barclay Undergrad Training/Plasmid Transformation/2013/10/03

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Current revision (17:17, 7 October 2013) (view source)
(Continuation of Restriction Digests and Gel Electrophoresis)
 
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| bgcolor=#cfcfcf | Reagent  
| bgcolor=#cfcfcf | Reagent  
| bgcolor=#cfcfcf | Volume
| bgcolor=#cfcfcf | Volume
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| rowspan="7" | <u>Expected:</u><br>1. hPCD = 186 bp, 4968 bp<br>2. Ubc = 368 bp, 4968 bp<br>
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| rowspan="7" | <u>Expected:</u><br>1. fshPCD = 186 bp, 4968 bp<br>
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| rowspan="7" | [[Image:IMAG0329.jpg|400px|Today's gel]]<br>15 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
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| rowspan="7" | [[Image:Gel DKB 10-3-13.jpg|400px|Today's gel]]<br>15 μL/lane; 1% agarose; [http://openwetware.org/wiki/Image:KAH_Fermentas_GeneRuler_1kbplus.jpg Ladder]
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| DNA(plasmid) || 5.0 μL
| DNA(plasmid) || 5.0 μL
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'''Conclusion'''
'''Conclusion'''
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There were definite gel lines at ~4900 and faint lines at ~200 for hPCD (lanes 1,2,5) and ~ 350 for Ubc (3,4,6). <br>
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There were faint bands at the 186 position; however, they are harder to see on the picture. The 4968 bands for all 3 are very visible. Fairly successful digest.
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This signifies a working gel.
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'''DNA Sequencing Samples'''
'''DNA Sequencing Samples'''
<!-- * signs create an automatically bulleted list. Replace 'date' with the date you submitted the samples -->
<!-- * signs create an automatically bulleted list. Replace 'date' with the date you submitted the samples -->
<!-- The Order Number is shown at the top of your sequencing order form. Record in the indicated spot below -->
<!-- The Order Number is shown at the top of your sequencing order form. Record in the indicated spot below -->
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* Submitted to DNASU on: '8/14/13'
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* Submitted to DNASU on: 10/7/2013
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* Order Number: 8043
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* Order Number: 8213
<!-- # signs create an automatically numbered list. Replace Plasmid 1 & 2 with the names of your plasmids. Replace 'name' with the appropriate name of each primer. -->
<!-- # signs create an automatically numbered list. Replace Plasmid 1 & 2 with the names of your plasmids. Replace 'name' with the appropriate name of each primer. -->
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# Plasmid 1 (hPCD) - Forward P001
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# Plasmid 1 (fshPCD) - Forward P001
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# Plasmid 1 (hPCD) - Reverse P002
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# Plasmid 1 (fshPCD) - Reverse P002
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# Plasmid 4 (Ubc) - Forward P001
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# Plasmid 4 (Ubc) - Reverse P002
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Current revision

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Continuation of Restriction Digests and Gel Electrophoresis

Restriction Digest Table

  • Checked plasmid minipreps with EcoRI/PstI digests
Reagent Volume Expected:
1. fshPCD = 186 bp, 4968 bp
Today's gel
15 μL/lane; 1% agarose; Ladder
DNA(plasmid) 5.0 μL
10X FD Buffer 1.5
EcoRI 1.0
PstI 1.0
dH2O 6.5
  15 μL --> 37°C/ 15 min.

Conclusion There were faint bands at the 186 position; however, they are harder to see on the picture. The 4968 bands for all 3 are very visible. Fairly successful digest.

DNA Sequencing Samples

  • Submitted to DNASU on: 10/7/2013
  • Order Number: 8213
  1. Plasmid 1 (fshPCD) - Forward P001
  2. Plasmid 1 (fshPCD) - Reverse P002



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