CTR:Notebook/PIRE/2014/07/09: Difference between revisions
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*Clean up | *Clean up | ||
*Sonication: 8 cycles of 15 seconds on, 90 seconds off, High Power | *Sonication: 8 cycles of 15 seconds on, 90 seconds off, High Power | ||
[[Image:Greenbuls1shearedgels.png|500px]]<br> | |||
Do not have have gel image for 2nd shearing attempt (4 additional cycles), but looked better than first. Because final attempt with additional 2 cycles did not look right (more large fragments), decided to continue with subsequent steps to avoid further sonication problems. | |||
*End Repair: NEBENDRP on John Block B | |||
*P2 Ligation: NEBLIGAT on John Block B | |||
<br> | |||
2014-08-01: | |||
*Size Selection: 40μL beads to remove large fragments (decreased amount to reduce loss of DNA), 25μL to remove small | |||
*PCR: NEBPCR on John Block B | |||
[[Image:Reprep-greenbuls1-pcr.JPG|400px]] 1μL template, 2μL 100bp ladder, 5μL PCR product<br> | |||
→Successful amplification! | |||
<br> | |||
*PicoGreen: 8.44 ng/μL | |||
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Revision as of 12:39, 1 August 2014
PIRE RAD Library Preps | <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page <html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> |
RAD Library Prep - Olive Sunbirds Plate 2, Little Greenbuls Plates 1 & 2
2μL of sheared DNA in each well
Troubleshooting
1) Pre-cleaned DNA (post-P1-ligation), 2) Cleaned DNA, 3) Sheared DNA
1) 1μL template, 2) 5μL PCR product
1&2) Sunbirds Plate 1, 3&4) Greenbuls Plate 1 (from re-sheared template) Re-prep Greenbuls Plate 1 using leftover DNA from plate2014-07-31:
2014-08-01:
1μL template, 2μL 100bp ladder, 5μL PCR product
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