Editing 1141 p8H-DNA-Pol-1

== '''Cloning of 1141 p8H-DNA-Pol-1'''  ==

'''Purpose:''' Cloning of 1141 p8H-DNA-Pol-1 

'''Date: '''July 23, 2012<br> 

'''Notebook:&nbsp;''' 

'''Page:''' 

'''Clone ID: '''1141 

'''Experimenters: '''Jennifer Truong 

== '''Results'''  ==

=== Cloning  ===

Date:&nbsp;July 27, 2012<br> 

[[Image:0724 Amp.JPG]]<br> 

See lane 5, expected size 2788 bp <br> 

PCR using MG1665 genomic DNA 

DNA-Pol1-Nde1-F<br>catgtaCATATGgttcagatcccccaaaatc<br>DNA-Pol1-Sal1-R<br>(cgaaaactgggatcaggcgcac GTC GAC tca cgat)<br>atcgtgaGTCGACGCGGCCGCCgtgcgcctgatcccagttttcg<br> 

Digest into vector 1135 with RE&nbsp;Nde1 and Sal1 

[[Image:0726 Digestion.JPG]]<br> 

See lane 10 (insert)&nbsp;and lane 11 (vector) 

Cut out bands and ligate together and transform 

<br> 

Date:&nbsp;August 2, 2012<br> 

Screening came back bad, started all over again<br> 

[[Image:0802 Amp.JPG]]<br> 

See lane 4, expected size 2788 bp 

PCR using MG1665 genomic DNA 

DNA-Pol1-Nde1-F<br>catgtaCATATGgttcagatcccccaaaatc<br>DNA-Pol1-Sal1-R<br>(cgaaaactgggatcaggcgcac GTC GAC tca cgat)<br>atcgtgaGTCGACGCGGCCGCCgtgcgcctgatcccagttttcg 

1[[Image:1141 Digestion.JPG]] 

Ligate together, transform 

Digest into vector 1135 with RE Nde1 and Sal1 <br> 

=== <u></u>Colony Screening<br>  ===

Date: July 27, 2012 

[[Image:0727 Digestion 2.JPG]] 

Digested MiniPrep with RE Nde1 and Sal1, expected size 2788 bp 

Samples 2, 5 and 6 may be correct, submitted sample 5 for sequencing<br> 

Date:&nbsp;August 6, 2012<br> 

1[[Image:1141 0806 Screening.JPG]]<br> 

Digested MiniPrep with RE Nde1 and Sal1, expected size 2788 bp <br> 

No inserts 

<br> 

Date: Nov30, 2012 <br> 

[[Image:1141 1147 1277 1444PCR.JPG]]<br> 

Digest vector 1135 and 1141pcr products with Nde1 and Sal1 <br> 

[[Image:1141PCR 1135 1147PCR 1135DIGEST.JPG]] 

<br> Digested MiniPrep with RE Nde1 and Sal1, expected size 2788 bp 

[[Image:1141digest.JPG]]

== '''Result intepretation/Summary'''  ==

<br> 

== '''Conclusion'''  ==

<br> 

[[Category:Cloning]]