Stock No: 008320
Protocol 29081: Standard PCR Assay - Generic Cre Melt Curve Analysis
Version 2.3

Notes

Melting curve analysis is done using a Roche Light Cycler 480.

This assay will NOT distinguish hemizygous from homozygous transgenic animals.
The genotyping protocol(s) presented here have been optimized for reagents and conditions used by The Jackson Laboratory (JAX). To genotype animals, JAX recommends researchers validate the assay independently upon receipt of animals into their facility. Reaction cycling temperature and times may require additional optimization based on the specific genotyping reagents used.

Expected Results

Transgene Tm = 83°C +/- 1.0°C
Internal positive control Tm = 86.5°C+/- 1.0°C

 

Transgene = ~100 bp
Internal positive control = 324 bp

 

JAX Protocol

Protocol Primers

Primer 5' Label Sequence 5' → 3' 3' Label Primer Type Reaction Note
oIMR1084 GCG GTC TGG CAG TAA AAA CTA TC Transgene Forward A Cre F
oIMR1085 GTG AAA CAG CAT TGC TGT CAC TT Transgene Reverse A Cre R
oIMR7338 CTA GGC CAC AGA ATT GAA AGA TCT Internal Positive Control Forward A
oIMR7339 GTA GGT GGA AAT TCT AGC ATC ATC C Internal Positive Control Reverse A

Reaction A

Component Final Concentration
ddH2O
Kapa 2G HS buffer 1.30 X
MgCl2 2.60 mM
dNTP KAPA 0.26 mM
oIMR1084 0.50 uM
oIMR1085 0.50 uM
oIMR7338 0.50 uM
oIMR7339 0.50 uM
Glycerol 6.50 %
Dye 1.00 X
Kapa 2G HS taq polym 0.03 U/ul
DNA

Cycling

Step Temp °C Time Note
1 94.0 --
2 94.0 --
3 65.0 -- -0.5 C per cycle decrease
4 68.0 --
5 -- repeat steps 2-4 for 10 cycles (Touchdown)
6 94.0 --
7 60.0 --
8 72.0 --
9 -- repeat steps 6-8 for 28 cycles
10 72.0 --
11 10.0 -- hold
JAX uses a very high speed Taq (~1000 bp/sec), use cycling times recommended for your reagents.
JAX uses a 'touchdown' cycling protocol and therefore has not calculated the optimal annealing temperature for each set of primers.

Strains Using This Protocol

Stock Number Strain Name
005989 129;FVB-Tg(PTH-cre)4167Slib/J
017789 129S-Slc11a2tm2Nca/J
007179 129S.Cg-Ndor1Tg(UBC-cre/ERT2)1Ejb/J
007915 129S.FVB-Tg(Amh-cre)8815Reb/J
008523 129S6.Cg-Tg(NPHS2-cre)295Lbh/BroJ
009575 B6(129S4)-Et(cre/ERT2)119Rdav/J
009580 B6(129S4)-Et(cre/ERT2)1382Rdav/J
009581 B6(129S4)-Et(cre/ERT2)1642Rdav/J
009582 B6(129S4)-Et(cre/ERT2)1645Rdav/J
009583 B6(129S4)-Et(cre/ERT2)1957Rdav/J
009584 B6(129S4)-Et(cre/ERT2)2007Rdav/J
009585 B6(129S4)-Et(cre/ERT2)2047Rdav/J
009574 B6(129S4)-Et(cre/ERT2)21Rdav/J
009577 B6(129S4)-Et(cre/ERT2)296Rdav/J
009578 B6(129S4)-Et(cre/ERT2)398Rdav/J
009573 B6(129S4)-Et(cre/ERT2)4Rdav/J
009579 B6(129S4)-Et(cre/ERT2)837Rdav/J
022356 B6(129X1)-Tg(Cd4-cre/ERT2)11Gnri/J
017889 B6(Cg)-Shank3tm1.1Bux/J
021012 B6(D2)-Tg(CAG-Brainbow1.0)3Eggn/J
018057 B6.129(Cg)-Cul7tm2.1Jdec/J
008320 B6.129(Cg)-Leprtm2(cre)Rck/J
018445 B6.129-Mre11atm1.1Rchd/J
005697 B6.129-Otx1tm4(cre)Asim/J
021160 B6.129P2(Cg)-Cx3cr1tm2.1(cre/ERT2)Litt/WganJ
018054 B6.129P2(Cg)-Pitx2tm1.1Dmm/J
023726 B6.129P2-C1qatm1.1Tenn/J
034597 B6.129P2-Kif13aGt(AF0553)Wtsi/AhcJ
016934 B6.129P2-Lgr6tm2.1(cre/ERT2)Cle/J
018997 B6.129S(FVB)-Lhx3tm1.1Sjr/J
006600 B6.129S1-Mnx1tm4(cre)Tmj/J
016132 B6.129S4-Perptm2Att/J
006447 B6.129S6(CBA)-Cebpatm1Dgt/J
012900 B6.129S6-Stat5btm1Mam Stat5atm2Mam/Mmjax
006054 B6.C-Tg(CMV-cre)1Cgn/J
006230 B6.Cg-Cebpatm1Dgt Tg(Mx1-cre)1Cgn/J
009107 B6.Cg-H2az2Tg(Wnt1-cre)11Rth Tg(Wnt1-GAL4)11Rth/J
022865 B6.Cg-Trib2tm1.1(cre/ERT2)Hze/J
017346 B6.Cg-Tg(A930038C07Rik-cre)1Aibs/J
033345 B6.Cg-Tg(Cdh5-COP4*L132C,-lacZ)B28-2Mik/J
032889 B6.Cg-Tg(Cdh5-RHO/Adrb2,-lacZ)B27-3Mik/J
009352 B6.Cg-Tg(CDX2-cre*)189Erf/J
006368 B6.Cg-Tg(Cr2-cre)3Cgn/J
006663 B6.Cg-Tg(Eno2-cre)39Jme/J
009642 B6.Cg-Tg(GH1-cre)1Sac/J
003573 B6.Cg-Tg(Ins2-cre)25Mgn/J
003967 B6.Cg-Tg(Rbp3-cre)528Jxm/J
008863 B6.Cg-Tg(Tek-cre)1Ywa/J
012328 B6.Cg-Tg(Tyr-cre/ERT2)13Bos/J
006451 B6.FVB(129X1)-Tg(Sim1-cre)1Lowl/J
006137 B6.FVB-Tg(Cdh5-cre)7Mlia/J
011069 B6.FVB-Tg(Gh1-cre)bKnmn/J
003394 B6.FVB-Tg(Zp3-cre)3Mrt/J
021961 B6;129-Abcg2tm3.1(cre/ERT2)Bsor/J
017601 B6;129-Eif2s1tm1Rjk/J
024767 B6;129-Fostm1Mxu/Mmjax
016210 B6;129-Lrrk2tm3.1Shn/J
018319 B6;129-Plxnd1tm1.1Tmj/J
009600 B6;129-Six2tm3(EGFP/cre/ERT2)Amc/J
018773 B6;129-Sult2b1tm1Rus/J
022864 B6;129S-Rasgrf2tm1(cre/folA)Hze/J
017593 B6;129S-Sox2tm1(cre/ERT2)Hoch/J
009388 B6;129S1-Osr2tm2(cre)Jian/J
018019 B6;129S4-Srsf2tm1Xdfu/J
009576 B6;129S4-Et(cre/ERT2)278Rdav/J
017495 B6;129S7-Crim1tm1(GFP/cre/ERT2)Pzg/J
018937 B6;C-Tg(Erv4-cre/ERT2*)657Slp/J
009616 B6;C3-Tg(A930038C07Rik-cre)4Aibs/J
008844 B6;C3-Tg(Ctgf-cre)2Aibs/J
008839 B6;C3-Tg(Cyp39a1-cre)1Aibs/J
009117 B6;C3-Tg(Cyp39a1-cre)7Aibs/J
009111 B6;C3-Tg(Scnn1a-cre)1Aibs/J
009112 B6;C3-Tg(Scnn1a-cre)2Aibs/J
009613 B6;C3-Tg(Scnn1a-cre)3Aibs/J
009103 B6;C3-Tg(Wfs1-cre/ERT2)3Aibs/J
024926 B6;D2-Tg(Fshr-cre)1Ldu/J
003466 B6;D2-Tg(Sycp1-cre)4Min/J
018402 B6;FVB-Nrn1tm1.2Ndiv/J
003734 B6;FVB-Tg(GZMB-cre)1Jcb/J
004426 B6;SJL-Tg(Cga-cre)3Sac/J
003554 B6;SJL-Tg(Col2a1-cre)1Bhr/J
017738 B6;SJL-Tg(Foxl1-cre)1Khk/J
005249 B6;SJL-Tg(Krt1-15-cre/PGR)22Cot/J
007252 B6Ei.129S4-Tg(Prm-cre)58Og/EiJ
034731 B6N(129S4)-Srd5a3tm1c(EUCOMM)Wtsi/J
017741 B6N.129(Cg)-Mecp2tm1.1Joez/J
017704 B6N.129-Pik3catm1Jjz/J
016881 B6N.Cg-Adgra2tm1.1Bstc/J
019102 B6N.Cg-Tg(CAG-cre/Esr1*)5Amc/CjDswJ
018966 B6N.Cg-Tg(Camk2a-cre)T29-1Stl/J
018965 B6N.Cg-Tg(Fabp4-cre)1Rev/J
018960 B6N.Cg-Tg(Ins2-cre)25Mgn/J
018964 B6N.Cg-Tg(KRT14-cre)1Amc/J
019103 B6N.Cg-Tg(Nes-cre)1Kln/CjDswJ
003465 BALB/c-Tg(CMV-cre)1Cgn/J
013756 C.129S-Batf3tm1Kmm/J
004126 C.Cg-Cd19tm1(cre)Cgn Ighb/J
005673 C.Cg-Tg(Mx1-cre)1Cgn/J
006244 C.Cg-Tg(tetO-cre)1Jaw/J
009155 C57BL/6-Cldn6tm1(cre)Dkwu/J
016097 C57BL/6-Tg(Car1-cre)5Flt/J
008870 C57BL/6-Tg(Hspa2-cre)1Eddy/J
024034 C57BL/6-Tg(Pmch-cre)1Rck/J
034448 C57BL/6J-Tlr9em1Ldm/J
032080 C57BL/6J-Tg(Cd8a*-cre)B8Asin/J
018899 C57BL/6J-Tg(Krt6,-cre,-Cerulean)5Grsr/Grsr
021582 C57BL/6J-Tg(Mchr1-cre)1Emf/J
022883 C57BL/6J-Tg(Six6-cre)3Grsr/GrsrJ
023453 C57BL/6N-Lhx8tm1(cre,Cerulean)Grsr/2GrsrJ
012686 C57BL/6N-Tg(Ppp1r2-cre)4127Nkza/J
004600 FVB-Tg(GFAP-cre)25Mes/J
006139 FVB.Cg-Tg(ACTA1-cre)79Jme/J
006297 FVB.Cg-Tg(Eno2-cre)39Jme/J
008244 FVB.Cg-Tg(tetO-cre)1Jaw/J
003377 FVB/N-Tg(Zp3-cre)3Mrt/J
017678 FVB;129-Pink1tm1Aub Tg(Prnp-SNCA*A53T)AAub/J
023325 FVB;B6-Tg(Pbsn-cre)20Fwan/J
034795 NOD.Cg-Tg(CAG-cre/Esr1*)5Amc/J
013251 NOD.FVB-Tg(EIIa-cre)C5379Lmgd/J
004986 NOD/ShiLt-Tg(Ins2-cre)3Lt/LtJ
003855 NOD/ShiLt-Tg(Ins2-cre)5Lt/LtJ
035185 STOCK Adgrl1tm2c(EUCOMM)Hmgu/SudJ
018473 STOCK Egfrtm1.1Tyj/J
007916 STOCK En1tm2(cre)Wrst/J
007913 STOCK Gli1tm3(cre/ERT2)Alj/J
008783 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm3(SMN2/Smn1)Mrph Tg(CAG-cre/Esr1*)5Amc Tg(SMN2*delta7)4299Ahmb/J
022793 STOCK Gt(ROSA)26Sortm1(LRRK2*R1441C)Djmo/J
003829 STOCK H2az2Tg(Wnt1-cre)11Rth Tg(Wnt1-GAL4)11Rth/J
017298 STOCK Hmgn5tm1.1Mbus/J
032429 STOCK Krastm4Tyj Trp53tm1Brn Tg(Pdx1-cre/Esr1*)#Dam/J
018418 STOCK Lrig1tm1.1(cre/ERT2)Rjc/J
016945 STOCK Mir138-1tm1Mtm/Mmjax
004192 STOCK Mttptm2Sgy Ldlrtm1Her Apobtm2Sgy Tg(Mx1-cre)1Cgn/J
035045 STOCK Oprk1tm1.1(cre)Sros/J
022972 STOCK Pros1tm1Grl/J
019378 STOCK Ptf1atm2(cre/ESR1)Cvw/J
013086 STOCK Rxratm1Krc/J
023405 STOCK Tfap2atm1Hsv/J
007684 STOCK Tg(Atoh1-cre/Esr1*)14Fsh/J
004453 STOCK Tg(CAG-cre/Esr1*)5Amc/J
009615 STOCK Tg(Cartpt-cre)1Aibs/J
017336 STOCK Tg(Cd4-cre)1Cwi/BfluJ
005105 STOCK Tg(Chx10-EGFP/cre,-ALPP)2Clc/J
008852 STOCK Tg(En2-cre)22Alj/J
005938 STOCK Tg(Eno2-cre)39Jme/J
022763 STOCK Tg(Eno2-cre/ERT2)1Pohlk/J
019380 STOCK Tg(IVL-cre/ERT2)1Blpn/J
008582 STOCK Tg(Kcnc2-Cre)K128Stl/LetJ
017948 STOCK Tg(KRT18-cre/ERT2)23Blpn/J
003553 STOCK Tg(MMTV-cre)4Mam/J
008847 STOCK Tg(Msx2-cre/ERT)27Alj/J
009074 STOCK Tg(Myh6-cre)1Jmk/J
009102 STOCK Tg(Nefh-cre)12Kul/J
005667 STOCK Tg(Neurog3-cre)C1Able/J
006207 STOCK Tg(Pcp2-cre)1Amc/J
009606 STOCK Tg(Six2-EGFP/cre)1Amc/J
032646 STOCK Tg(Spo11-cre)1Rsw/PecoJ
006224 STOCK Tg(tetO-cre)1Jaw/J
024240 STOCK Tg(Tnnt2-cre)5Blh/JiaoJ
008851 STOCK Tg(Wnt1-cre/ERT)1Alj/J
018281 STOCK Tg(Wnt7a-EGFP/cre)#Bhr/Mmjax
161 strains use this protocol