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

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(Autocreate 2013/10/03 Entry for Haynes_Lab:Notebook/David_Barclay_Undergrad_Training/Plasmid_Transformation)
 
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=Continuation of Restriction Digests and Gel Electrophoresis=
Restriction Digest Table<br>
* Checked plasmid minipreps with EcoRI/PstI digests


{| {{table}} border="1" cellspacing="3"
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<!-- The | symbols on the next two lines start new cells in the same row. This does the same thing as ||, but you have to use | on new lines to set formatting for cells. -->
<!-- bgcolor=#cfcfcf *colors* the *background* of the Reagent and Volume cells grey.  -->
<!-- The next two "rowspan=7" cells span  all 7 rows in the table so that they can fit the "Expected" list and a gel image. rowspan is how you create merged rows in Wiki code. -->
<!-- Replace Plasmid 1 and 2 with the names of your plasmids. Replace insert size and vector size with appropriate information for your plasmids. If you only have one Plasmid, delete all the text for Plasmid 2
<!-- After you upload your gel image to OWW, replace "GelImage.jpg" with the name of your image file -->
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|- valign="top"
| bgcolor=#cfcfcf | Reagent
| bgcolor=#cfcfcf | Volume
| rowspan="7" | <u>Expected:</u><br>1. hPCD = 186 bp, 4968 bp<br>2. Ubc = 368 bp, 4968 bp<br>
| 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]
|-
| DNA(plasmid) || 5.0 μL
|-
| 10X FD Buffer || 1.5
|-
| EcoRI || 1.0
|-
| PstI || 1.0
|-
| dH<sub>2</sub>O || 6.5
|-
| &nbsp; || 15 μL --> 37°C/ 15 min.
|}
'''Conclusion'''
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>
This signifies a working gel.
'''DNA Sequencing 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 -->
* Submitted to DNASU on: '8/14/13'
* Order Number: 8043
<!-- # 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. -->
# Plasmid 1 (hPCD) - Forward P001
# Plasmid 1 (hPCD) - Reverse P002
# Plasmid 4 (Ubc) - Forward P001
# Plasmid 4 (Ubc) - Reverse P002




Revision as of 18:41, 3 October 2013

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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. hPCD = 186 bp, 4968 bp
2. Ubc = 368 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 definite gel lines at ~4900 and faint lines at ~200 for hPCD (lanes 1,2,5) and ~ 350 for Ubc (3,4,6).
This signifies a working gel.

DNA Sequencing Samples

  • Submitted to DNASU on: '8/14/13'
  • Order Number: 8043
  1. Plasmid 1 (hPCD) - Forward P001
  2. Plasmid 1 (hPCD) - Reverse P002
  3. Plasmid 4 (Ubc) - Forward P001
  4. Plasmid 4 (Ubc) - Reverse P002



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