Inducible Cre (MCM)

Inducible expression or deletion has been widely used in biology. The fusion protein Cre and mutant estrogen receptor (Cre-ER) is sequestered in the cytosol. When encountered with Tamoxifen, Cre-ER is translocated to the nucleus and modify the genomic locus. In 1996, Dr. M Reth found that one more copy of the mutant estrogen receptor increases the inducibility and activity of Cre. In the fusion protein which they named mutant estrogen receptor Cre mutant estrogen receptor (MCM), the Cre molecule is flanked by two mutant estrogen receptor. The functionality of MCM has been used and examined in multiple tissues.

Ref: Zhang Y, et al Nucleic Acids Res. 1996.

MCM 5:

ACC TGC TCC TGG AGA TGT TG

MCM 3:

CAT CAC TCG TTG CAT CGA CC

Product: 273 bp.

p>Recommended condition:

94C 5 min
94C 30 sec
58C 30 sec
72C 30 sec
X 35 cycles
72C 5 min
4C -

Recommended reaction:

18.5 ul H2O
2.5 ul PCR buffer (10X)
0.5 ul dNTP (10 mM each)
0.5 ul MCM 5 (10 uM)
0.5 ul MCM 3 (10 uM)
0.5 ul Taq (5 U/ul)
2 ul DNA
--------
25 ul

These primers were designed and tested by Wang. These primers have been used in mouse tail DNA preparations to genotype MCM transgenic animals. These primers only pick up the transgene MCM, without crossing reaction with regular Cre.


Comments

Add a comment
Posted by Han Mon, 13 Jun 2011 21:25:40
Subject: Reference
Could you please provide complete information of the author who designed the MCM3 & 5 primers? Thank you.
Site News
New feature:
You can add comments on protocols now. Just go to any individual method page and click on the "Add a comment" link.

Highlight of the month
SIRT6 links glucose and lipid metabolism

SIRT1, a histone deacetylase, has drawn considerable attention due to its important roles in metabolic regulation and longevity. SIRT1 is induced by fasting and suppressed by feeding. SIRT1 increases glucose production and beta oxidation in liver to meet energy needs during food deprivation. Another member of SIRT family, SIRT6 follows similar expression patterns as SIRT1. The function of SIRT6 in metabolic regulation remains unknown.

Kim and colleagues found SIRT1 can regulate SIRT6 expression in liver. Specifically, SIRT1 functions together with FOXO3a and stimulates SIRT6 transcription during fasting. The activation of SIRT6 directly suppresses gene expression of metabolic enzymes involved in triglyceride synthesis and glycolysis in liver. Deficiency of SIRT6 causes increases of glucose utilization, reduced beta oxidation and as a result, fatty liver. More importantly, Kim et al found the expression of SIRT6 is decreased in human fatty liver samples which indicates SIRT6 may play critical roles in liver steatosis in clinical settings. This study landed in Cell Metabolism of this month. ... Read more highlights.