Protocol 29081: Standard PCR Assay - Generic Cre Melt Curve Analysis

STOCK Kras^tm4Tyj Trp53^tm1Brn Tg(Pdx1-cre/Esr1*)#Dam/J

Stock No: 032429

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-Slc11a2^tm2Nca/J
036336	129S.Cg-Ghrhr^tm1.1Deh Trhr^tm1.1Deh/Deh
007179	129S.Cg-Ndor1^Tg(UBC-cre/ERT2)1Ejb/J
007915	129S.FVB-Tg(Amh-cre)8815Reb/J
004302	129S1/Sv-Hprt1^tm1(CAG-cre)Mnn/J
008523	129S6.Cg-Tg(NPHS2-cre)295Lbh/BroJ
032051	B6(129S4)-Kit^tm1.1(cre)Jmol/J
037511	B6(129S4)-Pou2f3^tm1.1(cre/ERT2)Imt/J
024976	B6(129S4)-Tspo^tm1.1Maf/J
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
038514	B6(129S6)-Tg(Slc6a5-cre/ERT2)#Uze/J
039195	B6(Cg)-Il17a^tm1.1(EYFP/cre)Ehs/J
026512	B6(Cg)-Tg(TPO-cre/ERT2)1139Tyj/J
021012	B6(D2)-Tg(CAG-Brainbow1.0)3Eggn/J
037052	B6(FVB)-Tg(Sprr2f-cre)2Dcas/J
018057	B6.129(Cg)-Cul7^tm2.1Jdec/J
008320	B6.129(Cg)-Lepr^tm2(cre)Rck/J
033964	B6.129(Cg)-Ptf1a^tm2(cre/ESR1)Cvw Kras^tm4Tyj Pten^tm1Hwu/J
036564	B6.129(Cg)-Scn10a^tm2(cre)Jwo/TjpJ
008463	B6.129-Gt(ROSA)26Sor^tm1(cre/ERT2)Tyj/J
018445	B6.129-Mre11a^tm1.1Rchd/J
025567	B6.129-Olig2^tm1.1(cre)Wdr/J
005697	B6.129-Otx1^tm4(cre)Asim/J
004146	B6.129-Tg(Pcp2-cre)2Mpin/J
021160	B6.129P2(Cg)-Cx3cr1^tm2.1(cre/ERT2)Litt/WganJ
018054	B6.129P2(Cg)-Pitx2^tm1.1Dmm/J
023726	B6.129P2-C1qa^tm1.1Tenn/J
016934	B6.129P2-Lgr6^tm2.1(cre/ERT2)Cle/J
018997	B6.129S(FVB)-Lhx3^tm1.1Sjr/J
016132	B6.129S4-Perp^tm2Att/J
006447	B6.129S6(CBA)-Cebpa^tm1Dgt/J
012900	B6.129S6-Stat5b^tm1Mam Stat5a^tm2Mam/Mmjax
008601	B6.Cg-7630403G23Rik^Tg(Th-cre)1Tmd/J
006230	B6.Cg-Cebpa^tm1Dgt Tg(Mx1-cre)1Cgn/J
036450	B6.Cg-Foxl1^tm2(cre/ERT2)Khk/Mmjax
036337	B6.Cg-Ghrhr^tm1.1Deh Trhr^tm1.1Deh Tg(CAG-cre/Esr1*)5Amc/Deh
009107	B6.Cg-H2az2^Tg(Wnt1-cre)11Rth Tg(Wnt1-GAL4)11Rth/J
032923	B6.Cg-Kit^tm1(cre)Htng/J
028536	B6.Cg-Ndnf^tm1.1(folA/cre)Hze/J
030544	B6.Cg-Npy1r^tm1.1(cre/GFP)Rpa/J
031485	B6.Cg-Pou4f2^tm1(cre)Bnt/SjJ
038835	B6.Cg-Sdk1^em1(cre/GFP)Jrs/KrishJ
017889	B6.Cg-Shank3^tm1.1Bux/J
022865	B6.Cg-Trib2^tm1.1(cre/ERT2)Hze/J
017346	B6.Cg-Tg(A930038C07Rik-cre)1Aibs/J
006149	B6.Cg-Tg(ACTA1-cre)79Jme/J
030215	B6.Cg-Tg(Amigo2-cre)1Sieg/J
039983	B6.Cg-Tg(AR*121Q)3Als/J
009352	B6.Cg-Tg(CDX2-cre*)189Erf/J
006368	B6.Cg-Tg(Cr2-cre)3Cgn/J
029594	B6.Cg-Tg(Dmp1-cre/ERT2)D77Pdp/J
006663	B6.Cg-Tg(Eno2-cre)39Jme/J
035668	B6.Cg-Tg(Gata1-cre)1Sho/MdfJ
009642	B6.Cg-Tg(GH1-cre)1Sac/J
003573	B6.Cg-Tg(Ins2-cre)25Mgn/J
005584	B6.Cg-Tg(Prrx1-cre)1Cjt/J
032873	B6.Cg-Tg(Pthlh-cre/ERT2)909Nono/J
003967	B6.Cg-Tg(Rbp3-cre)528Jxm/J
021614	B6.Cg-Tg(S100A8-cre,-EGFP)1Ilw/J
004128	B6.Cg-Tg(Tek-cre)12Flv/J
008863	B6.Cg-Tg(Tek-cre)1Ywa/J
006234	B6.Cg-Tg(tetO-cre)1Jaw/J
012328	B6.Cg-Tg(Tyr-cre/ERT2)13Bos/J
035670	B6.Cg-Tg(VAV1-cre)1Graf/MdfJ
020282	B6.Cg-Tg(Vil1-cre/ERT2)23Syr/J
006451	B6.FVB(129X1)-Tg(Sim1-cre)1Lowl/J
006137	B6.FVB-Tg(Cdh5-cre)7Mlia/J
018980	B6.FVB-Tg(Ddx4-cre)1Dcas/KnwJ
003724	B6.FVB-Tg(EIIa-cre)C5379Lmgd/J
011069	B6.FVB-Tg(Gh1-cre)bKnmn/J
011038	B6.FVB-Tg(Myh6-cre)2182Mds/J
010714	B6.FVB-Tg(Pomc-cre)1Lowl/J
003394	B6.FVB-Tg(Zp3-cre)3Mrt/J
017601	B6;129-Eif2s1^tm1Rjk/J
016210	B6;129-Lrrk2^tm3.1Shn/J
018319	B6;129-Plxnd1^tm1.1Tmj/J
009600	B6;129-Six2^tm3(EGFP/cre/ERT2)Amc/J
018773	B6;129-Sult2b1^tm1Rus/J
022864	B6;129S-Rasgrf2^tm1(cre/folA)Hze/J
017593	B6;129S-Sox2^tm1(cre/ERT2)Hoch/J
009388	B6;129S1-Osr2^tm2(cre)Jian/J
036300	B6;129S4-Cux1^tm1(cre/ERT2)Zjh/J
018019	B6;129S4-Srsf2^tm1Xdfu/J
009576	B6;129S4-Et(cre/ERT2)278Rdav/J
017495	B6;129S7-Crim1^tm1(GFP/cre/ERT2)Pzg/J
018937	B6;C-Tg(Erv4-cre/ERT2*)657Slp/J
038989	B6;C3-Oxtr^em4(icre/ERT2)Yinn/2J
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
025809	B6;CBA-Tg(Fgfr3-icre/ERT2)4-2Wdr/J
024926	B6;D2-Tg(Fshr-cre)1Ldu/J
003466	B6;D2-Tg(Sycp1-cre)4Min/J
015855	B6;DBA-Tg(Upk3a-GFP/cre/ERT2)26Amc/J
018402	B6;FVB-Nrn1^tm1.2Ndiv/J
023748	B6;FVB-Tg(Aldh1l1-cre)JD1884Htz/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
003552	B6129-Tg(Wap-cre)11738Mam/J
007252	B6Ei.129S4-Tg(Prm-cre)58Og/EiJ
017741	B6N.129(Cg)-Mecp2^tm1.1Joez/J
017704	B6N.129-Pik3ca^tm1Jjz/J
016881	B6N.Cg-Adgra2^tm1.1Bstc/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-Batf3^tm1Kmm/J
004126	C.Cg-Cd19^tm1(cre)Cgn Igh^b/J
005673	C.Cg-Tg(Mx1-cre)1Cgn/J
006244	C.Cg-Tg(tetO-cre)1Jaw/J
009155	C57BL/6-Cldn6^tm1(cre)Dkwu/J
035489	C57BL/6-Elmo1^tm1Mki/J
038053	C57BL/6-Oprm1^tm2.1(EGFP/cre)Ics/KffJ
016097	C57BL/6-Tg(Car1-cre)5Flt/J
008870	C57BL/6-Tg(Hspa2-cre)1Eddy/J
024034	C57BL/6-Tg(Pmch-cre)1Rck/J
032131	C57BL/6-Tg(Scx-GFP/cre)1Stzr/J
036429	C57BL/6J-Gne^tm1Dar/J
039298	C57BL/6J-Shox2^em1(cre)Zyliu/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
027789	C57BL/6N-Fgd5^tm3(cre/ERT2)Djr/J
023453	C57BL/6N-Lhx8^tm1(cre)Grsr/2GrsrJ
012686	C57BL/6N-Tg(Ppp1r2-cre)4127Nkza/J
012929	FVB(Cg)-Tg(Dhh-cre)1Mejr/J
004600	FVB-Tg(GFAP-cre)25Mes/J
005025	FVB.Cg-Grm7^Tg(SMN2)89Ahmb Smn1^tm1Msd Tg(SMN2*delta7)4299Ahmb/J
006139	FVB.Cg-Tg(ACTA1-cre)79Jme/J
006297	FVB.Cg-Tg(Eno2-cre)39Jme/J
018394	FVB.Cg-Tg(KRT5-cre/ERT2)2Ipc/JeldJ
008244	FVB.Cg-Tg(tetO-cre)1Jaw/J
003314	FVB/N-Tg(EIIa-cre)C5379Lmgd/J
003377	FVB/N-Tg(Zp3-cre)3Mrt/J
017678	FVB;129-Pink1^tm1Aub Tg(Prnp-SNCA*A53T)AAub/J
023325	FVB;B6-Tg(Pbsn-cre)20Fwan/J
034859	NOD.Cg-Prkdc^scid Il2rg^tm1Wjl Tg(CAG-cre/Esr1*)5Amc/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 Adgrl1^tm2c(EUCOMM)Hmgu/SudJ
036942	STOCK Ager^tm2.1(cre/ERT2)Blh/2J
039535	STOCK Aldh1a3^tm1(cre/ERT2)Dac/J
037028	STOCK Calcr^tm1.1(cre)Mgmj/J
007916	STOCK En1^tm2(cre)Wrst/J
007913	STOCK Gli1^tm3(cre/ERT2)Alj/J
008783	STOCK Grm7^Tg(SMN2)89Ahmb Smn1^tm3(SMN2/Smn1)Mrph Tg(CAG-cre/Esr1*)5Amc Tg(SMN2*delta7)4299Ahmb/J
022793	STOCK Gt(ROSA)26Sor^tm1(LRRK2*R1441C)Djmo/J
003829	STOCK H2az2^Tg(Wnt1-cre)11Rth Tg(Wnt1-GAL4)11Rth/J
017298	STOCK Hmgn5^tm1.1Mbus/J
035978	STOCK Hoxb8^tm2.1(cre)Mrc/J
032429	STOCK Kras^tm4Tyj Trp53^tm1Brn Tg(Pdx1-cre/Esr1*)#Dam/J
018418	STOCK Lrig1^tm1.1(cre/ERT2)Rjc/J
016945	STOCK Mir138-1^tm1Mtm/Mmjax
004192	STOCK Mttp^tm2Sgy Ldlr^tm1Her Apob^tm2Sgy Tg(Mx1-cre)1Cgn/J
037139	STOCK Myl1^tm1(cre)Sjb Atg7^tm1Tchi Gaa^tm1Rabn/PuroMmjax
035303	STOCK Obscn^tm1Chen/J
035925	STOCK Opn4^tm1(cre)Saha/J
035045	STOCK Oprk1^tm1.1(cre)Sros/J
030543	STOCK Oxtr^tm1(cre/GFP)Rpa/J
022972	STOCK Pros1^tm1Grl/J
019378	STOCK Ptf1a^tm2(cre/ESR1)Cvw/J
013086	STOCK Rxra^tm1Krc/J
020080	STOCK Slc6a3^tm1(cre)Xz/J
023405	STOCK Tfap2a^tm1Hsv/J
036906	STOCK Tmem178b^tm1c(KOMP)Wtsi Tmem178^tm1.1Sud/J
010911	STOCK Wt1^tm1(EGFP/cre)Wtp/J
007684	STOCK Tg(Atoh1-cre/Esr1*)14Fsh/J
004453	STOCK Tg(CAG-cre/Esr1*)5Amc/J
035566	STOCK Tg(CAG-mRuby2*,-mClover*)1Srgt/J
009615	STOCK Tg(Cartpt-cre)1Aibs/J
017336	STOCK Tg(Cd4-cre)1Cwi/BfluJ
025736	STOCK Tg(Cela1-cre/ERT)1Lgdn/J
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
027681	STOCK Tg(Hnf1b-cre/ERT2)1Jfer/J
019380	STOCK Tg(IVL-cre/ERT2)1Blpn/J
008582	STOCK Tg(Kcnc2-Cre)K128Stl/LetJ
005107	STOCK Tg(KRT14-cre/ERT)20Efu/J
017948	STOCK Tg(KRT18-cre/ERT2)23Blpn/J
009643	STOCK Tg(Lhb-cre)1Sac/J
023724	STOCK Tg(mI56i-cre,EGFP)1Kc/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
028364	STOCK Tg(Neurod1-cre)1Able/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
223 strains use this protocol