Genotyping Protocols Database
Stock No: 008870
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 = 84°C +/- 1.0°C
Internal positive control Tm = 79.4°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
003377 FVB/N-Tg(Zp3-cre)3Mrt/J
003394 B6.FVB-Tg(Zp3-cre)3Mrt/J
003465 BALB/c-Tg(CMV-cre)1Cgn/J
003466 B6;D2-Tg(Sycp1-cre)4Min/J
003554 B6;SJL-Tg(Col2a1-cre)1Bhr/J
003734 B6;FVB-Tg(GZMB-cre)1Jcb/J
003829 STOCK H2az2Tg(Wnt1-cre)11Rth Tg(Wnt1-GAL4)11Rth/J
003855 NOD/ShiLt-Tg(Ins2-cre)5Lt/LtJ
003967 B6.Cg-Tg(Rbp3-cre)528Jxm/J
004192 STOCK Mttptm2Sgy Ldlrtm1Her Apobtm2Sgy Tg(Mx1-cre)1Cgn/J
004426 B6;SJL-Tg(Cga-cre)3Sac/J
004453 STOCK Tg(CAG-cre/Esr1*)5Amc/J
004600 FVB-Tg(GFAP-cre)25Mes/J
004986 NOD/ShiLt-Tg(Ins2-cre)3Lt/LtJ
005105 STOCK Tg(Chx10-EGFP/cre,-ALPP)2Clc/J
005249 B6;SJL-Tg(Krt1-15-cre/PGR)22Cot/J
005667 STOCK Tg(Neurog3-cre)C1Able/J
005673 C.Cg-Tg(Mx1-cre)1Cgn/J
005697 B6.129-Otx1tm4(cre)Asim/J
005938 STOCK Tg(Eno2-cre)39Jme/J
005989 129;FVB-Tg(PTH-cre)4167Slib/J
006054 B6.C-Tg(CMV-cre)1Cgn/J
006137 B6.FVB-Tg(Cdh5-cre)7Mlia/J
006139 FVB.Cg-Tg(ACTA1-cre)79Jme/J
006207 STOCK Tg(Pcp2-cre)1Amc/J
006224 STOCK Tg(tetO-cre)1Jaw/J
006230 B6.Cg-Cebpatm1Dgt Tg(Mx1-cre)1Cgn/J
006244 C.Cg-Tg(tetO-cre)1Jaw/J
006297 FVB.Cg-Tg(Eno2-cre)39Jme/J
006368 B6.Cg-Tg(Cr2-cre)3Cgn/J
006395 STOCK Tg(Sim1-cre)1Lowl/J
006447 B6.129S6(CBA)-Cebpatm1Dgt/J
006451 B6.FVB(129X1)-Tg(Sim1-cre)1Lowl/J
006600 B6.129S1-Mnx1tm4(cre)Tmj/J
006663 B6.Cg-Tg(Eno2-cre)39Jme/J
007179 129S.Cg-Ndor1Tg(UBC-cre/ERT2)1Ejb/J
007252 B6Ei.129S4-Tg(Prm-cre)58Og/EiJ
007684 STOCK Tg(Atoh1-cre/Esr1*)14Fsh/J
007913 STOCK Gli1tm3(cre/ERT2)Alj/J
007915 129S.FVB-Tg(Amh-cre)8815Reb/J
007916 STOCK En1tm2(cre)Wrst/J
008244 FVB.Cg-Tg(tetO-cre)1Jaw/J
008320 B6.129(Cg)-Leprtm2(cre)Rck/J
008523 129S6.Cg-Tg(NPHS2-cre)295Lbh/BroJ
008582 STOCK Tg(Kcnc2-Cre)K128Stl/LetJ
008783 STOCK Grm7Tg(SMN2)89Ahmb Smn1tm3(SMN2/Smn1)Mrph Tg(CAG-cre/Esr1*)5Amc Tg(SMN2*delta7)4299Ahmb/J
008839 B6;C3-Tg(Cyp39a1-cre)1Aibs/J
008844 B6;C3-Tg(Ctgf-cre)2Aibs/J
008847 STOCK Tg(Msx2-cre/ERT)27Alj/J
008851 STOCK Tg(Wnt1-cre/ERT)1Alj/J
008852 STOCK Tg(En2-cre)22Alj/J
008870 C57BL/6-Tg(Hspa2-cre)1Eddy/J
009074 STOCK Tg(Myh6-cre)1Jmk/J
009102 STOCK Tg(Nefh-cre)12Kul/J
009103 B6;C3-Tg(Wfs1-cre/ERT2)3Aibs/J
009107 B6.Cg-H2az2Tg(Wnt1-cre)11Rth Tg(Wnt1-GAL4)11Rth/J
009111 B6;C3-Tg(Scnn1a-cre)1Aibs/J
009112 B6;C3-Tg(Scnn1a-cre)2Aibs/J
009117 B6;C3-Tg(Cyp39a1-cre)7Aibs/J
009155 C57BL/6-Cldn6tm1(cre)Dkwu/J
009352 B6.Cg-Tg(CDX2-cre*)189Erf/J
009388 B6;129S1-Osr2tm2(cre)Jian/J
009573 B6(129S4)-Et(cre/ERT2)4Rdav/J
009574 B6(129S4)-Et(cre/ERT2)21Rdav/J
009575 B6(129S4)-Et(cre/ERT2)119Rdav/J
009576 B6;129S4-Et(cre/ERT2)278Rdav/J
009577 B6(129S4)-Et(cre/ERT2)296Rdav/J
009578 B6(129S4)-Et(cre/ERT2)398Rdav/J
009579 B6(129S4)-Et(cre/ERT2)837Rdav/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
009600 B6;129-Six2tm3(EGFP/cre/ERT2)Amc/J
009606 STOCK Tg(Six2-EGFP/cre)1Amc/J
009613 B6;C3-Tg(Scnn1a-cre)3Aibs/J
009614 B6.Cg-Tg(Wfs1-cre/ERT2)2Aibs/J
009615 STOCK Tg(Cartpt-cre)1Aibs/J
009616 B6;C3-Tg(A930038C07Rik-cre)4Aibs/J
009642 B6.Cg-Tg(GH1-cre)1Sac/J
010708 B6(Cg)-Ssttm1(cre/ERT2)Zjh/J
011069 B6.FVB-Tg(Gh1-cre)bKnmn/J
012328 B6.Cg-Tg(Tyr-cre/ERT2)13Bos/J
012686 C57BL/6N-Tg(Ppp1r2-cre)4127Nkza/J
012900 B6.129S6-Stat5btm1Mam Stat5atm2Mam/Mmjax
013086 STOCK Rxratm1Krc/J
013251 NOD.FVB-Tg(EIIa-cre)C5379Lmgd/J
013756 C.129S-Batf3tm1Kmm/J
016097 C57BL/6-Tg(Car1-cre)5Flt/J
016132 B6.129S4-Perptm2Att/J
016210 B6;129-Lrrk2tm3.1Shn/J
016881 B6N.Cg-Adgra2tm1.1Bstc/J
016934 B6.129P2-Lgr6tm2.1(cre/ERT2)Cle/J
016945 STOCK Mir138-1tm1Mtm/Mmjax
016974 FVB;129P2-Mettm1Sst/J
017298 STOCK Hmgn5tm1.1Mbus/J
017336 STOCK Tg(Cd4-cre)1Cwi/BfluJ
017346 B6.Cg-Tg(A930038C07Rik-cre)1Aibs/J
017593 B6;129S-Sox2tm1(cre/ERT2)Hoch/J
017601 B6;129-Eif2s1tm1Rjk/J
017678 FVB;129-Pink1tm1Aub Tg(Prnp-SNCA*A53T)AAub/J
017704 B6N.129-Pik3catm1Jjz/J
017738 B6;SJL-Tg(Foxl1-cre)1Khk/J
017741 B6N.129(Cg)-Mecp2tm1.1Joez/J
017789 129S-Slc11a2tm2Nca/J
017889 B6(Cg)-Shank3tm1.1Bux/J
017948 STOCK Tg(KRT18-cre/ERT2)23Blpn/J
018019 B6;129S4-Srsf2tm1Xdfu/J
018054 B6.129P2(Cg)-Pitx2tm1.1Dmm/J
018057 B6.129(Cg)-Cul7tm2.1Jdec/J
018281 STOCK Tg(Wnt7a-EGFP/cre)#Bhr/Mmjax
018319 B6;129-Plxnd1tm1.1Tmj/J
018402 B6;FVB-Nrn1tm1.2Ndiv
018418 STOCK Lrig1tm1.1(cre/ERT2)Rjc/J
018445 B6.129-Mre11atm1.1Rchd/J
018473 STOCK Egfrtm1.1Tyj/J
018773 B6;129-Sult2b1tm1Rus/J
018899 C57BL/6J-Tg(Krt6,-cre,-Cerulean)5Grsr/Grsr
018937 B6;C-Tg(Erv4-cre/ERT2*)657Slp/J
018960 B6N.Cg-Tg(Ins2-cre)25Mgn/J
018965 B6N.Cg-Tg(Fabp4-cre)1Rev/J
018966 B6N.Cg-Tg(Camk2a-cre)T29-1Stl/J
018997 B6.129S(FVB)-Lhx3tm1.1Sjr/J
019102 B6N.Cg-Tg(CAG-cre/Esr1*)5Amc/CjDswJ
019103 B6N.Cg-Tg(Nes-cre)1Kln/CjDswJ
019378 STOCK Ptf1atm2(cre/ESR1)Cvw/J
019380 STOCK Tg(IVL-cre/ERT2)1Blpn/J
021012 B6(D2)-Tg(CAG-Brainbow1.0)3Eggn/J
021160 B6.129P2(Cg)-Cx3cr1tm2.1(cre/ERT2)Litt/WganJ
021582 C57BL/6J-Tg(Mchr1-cre)1Emf/J
021961 B6;129-Abcg2tm3.1(cre/ERT2)Bsor/J
022356 B6(129X1)-Tg(Cd4-cre/ERT2)11Gnri/J
022763 STOCK Tg(Eno2-cre/ERT2)1Pohlk/J
022793 STOCK Gt(ROSA)26Sortm1(LRRK2*R1441C)Djmo/J
022864 B6;129S-Rasgrf2tm1(cre/folA)Hze/J
022883 C57BL/6J-Tg(Six6-cre)3Grsr/GrsrJ
022972 STOCK Pros1tm1Grl/J
023325 FVB;B6-Tg(Pbsn-cre)20Fwan/J
023405 STOCK Tfap2atm1Hsv/J
023453 C57BL/6N-Lhx8tm1(cre,Cerulean)Grsr/2GrsrJ
023726 B6.129P2-C1qatm1.1Tenn/J
024034 C57BL/6-Tg(Pmch-cre)1Rck/J
024240 STOCK Tg(Tnnt2-cre)5Blh/JiaoJ
024767 B6;129-Fostm1Mxu/Mmjax
024926 B6;D2-Tg(Fshr-cre)1Ldu/J
147 strains use this protocol