User:Ajeffs/Notebook/Melanoma/2008/05/19: Difference between revisions

From OpenWetWare
< User:Ajeffs‎ | Notebook‎ | Melanoma‎ | 2008‎ | 05
Jump to navigationJump to search
Line 8: Line 8:
===siRNA-mediated knockdown of PAX3 and MITF in NZM12 and NZM15===
===siRNA-mediated knockdown of PAX3 and MITF in NZM12 and NZM15===
*Reverse transfection with Dharmacon Smartpools (10 nM MITF; 20 nM PAX3; 20 nM non-targeting pool).
*Reverse transfection with Dharmacon Smartpools (10 nM MITF; 20 nM PAX3; 20 nM non-targeting pool).
**Assess transcript knock-down at 24 hours post-txn by using qPCR
*#Assess transcript knock-down at 24 hours post-txn by using qPCR
**Assess protein levels 48 hours post-txn by using Western
*#Assess protein levels 48 hours post-txn by using Western
**n = 2 of 3 replicate experiments.
*#n = 2 of 3 replicate experiments.
*Measure EDNRB, RAB27A, FOXD3, and SNAI2 transcript levels by using qPCR at 24 and 48 hours post-txn.
*Measure EDNRB, RAB27A, FOXD3, and SNAI2 transcript levels by using qPCR at 24 and 48 hours post-txn.
===Does miR-137 repeat expansion correlate with MITF levels in NZM cells?===
===Does miR-137 repeat expansion correlate with MITF levels in NZM cells?===
*Assess miR-137 locus stability at 1p22 with conventional PCR.
*Assess miR-137 locus stability at 1p22 with conventional PCR.

Revision as of 17:26, 18 May 2008

Kaloyan's melanoma project <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>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>

The effect of PAX3 and MITF knock-down in melanoma cell lines

siRNA-mediated knockdown of PAX3 and MITF in NZM12 and NZM15

  • Reverse transfection with Dharmacon Smartpools (10 nM MITF; 20 nM PAX3; 20 nM non-targeting pool).
    1. Assess transcript knock-down at 24 hours post-txn by using qPCR
    2. Assess protein levels 48 hours post-txn by using Western
    3. n = 2 of 3 replicate experiments.
  • Measure EDNRB, RAB27A, FOXD3, and SNAI2 transcript levels by using qPCR at 24 and 48 hours post-txn.

Does miR-137 repeat expansion correlate with MITF levels in NZM cells?

  • Assess miR-137 locus stability at 1p22 with conventional PCR.
    1. purify and sequence a selection of the different sized amplicons.
    2. predict impact of repeat expansion on pre-miR folding.
    3. measure miR-137 VNTR status in non-melanoma/skin-derived cells?


Retrieved from "https://openwetware.org/mediawiki/index.php?title=User:Ajeffs/Notebook/Melanoma/2008/05/19&oldid=206121"