https://eln.excellgen.com/index.php?action=history&feed=atom&title=1439_pFus_B10-SMN2-L2
1439 pFus B10-SMN2-L2 - Revision history
2026-10-18T06:12:21Z
Revision history for this page on the wiki
MediaWiki 1.44.0
https://eln.excellgen.com/index.php?title=1439_pFus_B10-SMN2-L2&diff=7148&oldid=prev
Paul.xu at 21:02, 20 November 2012
2012-11-20T21:02:38Z
<p></p>
<p><b>New page</b></p><div>== '''Cloning of 1439 pFus B10-SMN2-L2''' ==<br />
<br />
'''Purpose:''' Cloning of 1439 pFus B10-SMN2-L2<br />
<br />
'''Date: '''2012-1--- <br />
<br />
'''Notebook:0''' <br />
<br />
'''Page:''' <br />
<br />
'''Clone ID:1439''' <br />
<br />
'''Experimenters:'''&nbsp; shanyi <br />
<br />
== '''Results''' ==<br />
<br />
Day2, pick white clones to PCR using primers pCR8_F1 and pCR8_R1&nbsp;: <br />
<br />
[[Image:SMN2---1 TOP1 1-8 and TOP2 1-8.JPG]]<br> <br />
<br />
<br />
Day3<br> <br />
<br />
&nbsp;Miniprep the&nbsp; plasmids: Use enzymes AflII and XbaI to digest<br> <br />
<br />
[[Image:Smn2--smn1digest 11 02-2012.JPG]]<br> <br />
<br />
Top1-5,Top1-8,Bottom1-1, Bottom2-1,Bottom2-4,Bottom2-5 may be right.<br> <br />
<br />
<br> <br />
<br />
Transform TOP2 again.<br> <br />
<br />
Day4, pick white clones to PCR using primers pCR8_F1 and pCR8_R1&nbsp;: <br />
<br> <br />
[[Image:SMN2---1 TOP2 1-14pcr.JPG]] <br />
<br />
Day5<br> <br />
<br />
Mini prep the plasmids: Use enzymes AflII and XbaI to digest<br> <br />
<br />
[[Image:Smn2--smn1digest 11 05-2012.JPG]]<br> <br />
<br />
Top2-8,Top1-12,Top1-14 may be right.<br> <br />
<br />
<br> <br />
<br />
Sequence TOP1-5,BOTTOM1-1,BOTTOM2-1 and top2-8,top2-14 with primers pCR8_F1 and pCR8_R1.<br> <br />
<br />
TOP1-5,BOTTOM1-1,BOTTOM2-1 are right.<br> <br />
<br />
Top2-8,Top2-14 are empty vectors, so need to repeat step1.<br> <br />
<br />
<br> <br />
<br />
<br> <br />
<br />
TOP2 .<br />
<br><br />
Repeat reaction1,digest and ligate again,<br />
<br />
pick white clones to PCR using primers pCR8_F1 and pCR8_R1&nbsp;: <br />
<br />
[[Image:Smn2--1top2.JPG]]<br />
<br> <br />
<br />
TOP2<br />
CHANGE F4 pFUS-B-10, C1,A6,D2,D4, do reaction#1 again.<br />
pick white clones to PCR using primers pCR8_F1 and pCR8_R1&nbsp;: <br />
still no band(dna polymerase is not good)<br />
<br />
Mini prep the plasmids: Use enzymes AflII and XbaI to digest<br> <br />
<br />
<br />
[[Image:Top2digest with AfIII AND xbaI.JPG]]<br />
<br />
Sequence TOP2N-1 with primers pCR8_F1 and pCR8_R1.<br />
<br />
Top2n-1 is right.<br />
<br />
<br />
<br />
<br />
[[Category:Cloning]]</div>
Paul.xu