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Contents:In the table below, Illumina Nextera Handle sequences have been added and highlighted in bold. Primers for the on-target have been added for convenience. The table below is sorted by the CFD off-target score. Sites with very low CFD scores < 0.02 are unlikely to be cleaved, see our study Haeussler et al. 2016, Figure 2.
In the protocol by Matthew Canver, Harvard, two PCRs are run: one PCR to amplify the potential off-target, then a second PCR to extend the handles with Illumina barcodes. Please click here to download the protocol. Alternatively, you can have a look at Fu et al, 2014.
If a primer was not found, the reason is usually that the region around the off-target is too repetitive. To avoid unspecific primers, all repeats are masked for the primer design (not for off-target search). If you think that we should change the parameters here or should use different primer3 settings, please let us know.
| Name | Primer Sequence | Tm | CFD Score |
|---|---|---|---|
| ontarget_mm0_exon_5730457N03Rik/Evx1_chr6_52314048_F | Primer3: not found at this Tm | N.d. | 1.00 |
| ontarget_mm0_exon_5730457N03Rik/Evx1_chr6_52314048_R | Primer3: not found at this Tm | N.d. | 1.00 |
| mm4_intergenic_Olfr714|Olfr17_chr7_107075417_F | Primer3: not found at this Tm | N.d. | 0.44 |
| mm4_intergenic_Olfr714|Olfr17_chr7_107075417_R | Primer3: not found at this Tm | N.d. | 0.44 |
| mm4_intergenic_Gm13814|Gm13814/Gm13817_chr2_92506885_F | Primer3: not found at this Tm | N.d. | 0.36 |
| mm4_intergenic_Gm13814|Gm13814/Gm13817_chr2_92506885_R | Primer3: not found at this Tm | N.d. | 0.36 |
| mm4_intron_C2cd2_chr16_97909395_F | TCGTCGGCAGCGTCAGGCTTCGGTTGCTAATTCAC | 58.9 | 0.33 |
| mm4_intron_C2cd2_chr16_97909395_R | GTCTCGTGGGCTCGGAGGAGGGACAGAGATGCAGA | 59.6 | 0.33 |
| mm4_intergenic_Gm13364|Gm6587_chr2_15755662_F | Primer3: not found at this Tm | N.d. | 0.31 |
| mm4_intergenic_Gm13364|Gm6587_chr2_15755662_R | Primer3: not found at this Tm | N.d. | 0.31 |
| mm4_intergenic_Gm2694|4933402J07Rik_chr8_87537233_F | Primer3: not found at this Tm | N.d. | 0.31 |
| mm4_intergenic_Gm2694|4933402J07Rik_chr8_87537233_R | Primer3: not found at this Tm | N.d. | 0.31 |
| mm4_intron_Ntrk2_chr13_58955411_F | TCGTCGGCAGCGTCGCTTGTGTGTGCTGACATGG | 60.0 | 0.28 |
| mm4_intron_Ntrk2_chr13_58955411_R | GTCTCGTGGGCTCGGGCACTTGATAGCCACACCCT | 60.0 | 0.28 |
| mm4_intergenic_Nrbf2|Egr2_chr10_67425600_F | Primer3: not found at this Tm | N.d. | 0.24 |
| mm4_intergenic_Nrbf2|Egr2_chr10_67425600_R | Primer3: not found at this Tm | N.d. | 0.24 |
| mm4_intron_Tpcn2_chr7_145268346_F | Primer3: not found at this Tm | N.d. | 0.24 |
| mm4_intron_Tpcn2_chr7_145268346_R | Primer3: not found at this Tm | N.d. | 0.24 |
| mm4_intron_Asb1_chr1_91542337_F | TCGTCGGCAGCGTCGTGAACCTTGTGCTGCCTTG | 59.9 | 0.23 |
| mm4_intron_Asb1_chr1_91542337_R | GTCTCGTGGGCTCGGTCCGTTGAGCCCCTACAGAT | 60.3 | 0.23 |
| mm4_intergenic_Gm22618|Gm22976_chr5_51164578_F | Primer3: not found at this Tm | N.d. | 0.23 |
| mm4_intergenic_Gm22618|Gm22976_chr5_51164578_R | Primer3: not found at this Tm | N.d. | 0.23 |
| mm4_intron_Lphn3_chr5_81321036_F | TCGTCGGCAGCGTCTGTTTCCTCACCAAAGCAACA | 58.5 | 0.21 |
| mm4_intron_Lphn3_chr5_81321036_R | GTCTCGTGGGCTCGGGCCTTCCACACTGAGGTTCA | 59.8 | 0.21 |
| mm4_intergenic_Treml4|Treml2_chr17_48296450_F | Primer3: not found at this Tm | N.d. | 0.19 |
| mm4_intergenic_Treml4|Treml2_chr17_48296450_R | Primer3: not found at this Tm | N.d. | 0.19 |
| mm4_intergenic_Gm22729|Oxsr1_chr9_119317001_F | Primer3: not found at this Tm | N.d. | 0.18 |
| mm4_intergenic_Gm22729|Oxsr1_chr9_119317001_R | Primer3: not found at this Tm | N.d. | 0.18 |
| mm4_intergenic_Cmklr1|1700069L16Rik_chr5_113691557_F | Primer3: not found at this Tm | N.d. | 0.18 |
| mm4_intergenic_Cmklr1|1700069L16Rik_chr5_113691557_R | Primer3: not found at this Tm | N.d. | 0.18 |
| mm4_exon_Srrm3_chr5_135840667_F | Primer3: not found at this Tm | N.d. | 0.16 |
| mm4_exon_Srrm3_chr5_135840667_R | Primer3: not found at this Tm | N.d. | 0.16 |
| mm4_intergenic_Gm10051|Gm15627_chr5_133972024_F | Primer3: not found at this Tm | N.d. | 0.16 |
| mm4_intergenic_Gm10051|Gm15627_chr5_133972024_R | Primer3: not found at this Tm | N.d. | 0.16 |
| mm4_intron_Spire1_chr18_67531463_F | Primer3: not found at this Tm | N.d. | 0.14 |
| mm4_intron_Spire1_chr18_67531463_R | Primer3: not found at this Tm | N.d. | 0.14 |
| mm3_intergenic_Gm25148|Tifab_chr13_56171309_F | TCGTCGGCAGCGTCTAGATGGATGACCTGGGGCA | 60.0 | 0.14 |
| mm3_intergenic_Gm25148|Tifab_chr13_56171309_R | GTCTCGTGGGCTCGGCCAAACCAGCTGGGGCAC | 61.3 | 0.14 |
| mm4_intron_Cacna2d3_chr14_29674996_F | Primer3: not found at this Tm | N.d. | 0.13 |
| mm4_intron_Cacna2d3_chr14_29674996_R | Primer3: not found at this Tm | N.d. | 0.13 |
| mm4_intergenic_Pcdha2|Pcdhb1_chr18_37202147_F | Primer3: not found at this Tm | N.d. | 0.13 |
| mm4_intergenic_Pcdha2|Pcdhb1_chr18_37202147_R | Primer3: not found at this Tm | N.d. | 0.13 |
| mm4_intron_Nsun2_chr13_69621726_F | Primer3: not found at this Tm | N.d. | 0.13 |
| mm4_intron_Nsun2_chr13_69621726_R | Primer3: not found at this Tm | N.d. | 0.13 |
| mm4_intergenic_Nrbf2|Egr2_chr10_67464115_F | Primer3: not found at this Tm | N.d. | 0.12 |
| mm4_intergenic_Nrbf2|Egr2_chr10_67464115_R | Primer3: not found at this Tm | N.d. | 0.12 |
| mm4_intergenic_Adra2c|E130018O15Rik_chr5_35314570_F | TCGTCGGCAGCGTCCCCGAGGGATGGAATGATGG | 59.9 | 0.11 |
| mm4_intergenic_Adra2c|E130018O15Rik_chr5_35314570_R | GTCTCGTGGGCTCGGTGTTCTTAGCCCTGCAGTGG | 59.9 | 0.11 |
| mm4_intergenic_Irx1|Rpl9-ps4_chr13_72231918_F | TCGTCGGCAGCGTCTAAAAGCCCATCGTCCCCAC | 60.0 | 0.11 |
| mm4_intergenic_Irx1|Rpl9-ps4_chr13_72231918_R | GTCTCGTGGGCTCGGAGCATGCCTACAACACTCCC | 60.0 | 0.11 |
| mm4_intergenic_Gria4|Gm23811/Gria4_chr9_4443109_F | Primer3: not found at this Tm | N.d. | 0.10 |
| mm4_intergenic_Gria4|Gm23811/Gria4_chr9_4443109_R | Primer3: not found at this Tm | N.d. | 0.10 |
| mm4_exon_Ccdc38_chr10_93573994_F | Primer3: not found at this Tm | N.d. | 0.10 |
| mm4_exon_Ccdc38_chr10_93573994_R | Primer3: not found at this Tm | N.d. | 0.10 |
| mm3_intron_Setbp1_chr18_78869194_F | Primer3: not found at this Tm | N.d. | 0.10 |
| mm3_intron_Setbp1_chr18_78869194_R | Primer3: not found at this Tm | N.d. | 0.10 |
| mm3_intergenic_Gm23892|Gm25382_chr10_16928421_F | Primer3: not found at this Tm | N.d. | 0.10 |
| mm3_intergenic_Gm23892|Gm25382_chr10_16928421_R | Primer3: not found at this Tm | N.d. | 0.10 |
| mm4_intergenic_Gm11986|U7_chr11_7633200_F | Primer3: not found at this Tm | N.d. | 0.09 |
| mm4_intergenic_Gm11986|U7_chr11_7633200_R | Primer3: not found at this Tm | N.d. | 0.09 |
| mm4_intron_Trpc4_chr3_54172959_F | Primer3: not found at this Tm | N.d. | 0.09 |
| mm4_intron_Trpc4_chr3_54172959_R | Primer3: not found at this Tm | N.d. | 0.09 |
| mm4_exon_Chtf8/Has3_chr8_106886388_F | Primer3: not found at this Tm | N.d. | 0.07 |
| mm4_exon_Chtf8/Has3_chr8_106886388_R | Primer3: not found at this Tm | N.d. | 0.07 |
| mm4_intergenic_Gm8601|Btg1_chr10_96599750_F | Primer3: not found at this Tm | N.d. | 0.06 |
| mm4_intergenic_Gm8601|Btg1_chr10_96599750_R | Primer3: not found at this Tm | N.d. | 0.06 |
| mm4_intergenic_Fam227b|Fgf7_chr2_126032200_F | Primer3: not found at this Tm | N.d. | 0.05 |
| mm4_intergenic_Fam227b|Fgf7_chr2_126032200_R | Primer3: not found at this Tm | N.d. | 0.05 |
| mm4_intron_Jakmip1_chr5_37146267_F | Primer3: not found at this Tm | N.d. | 0.05 |
| mm4_intron_Jakmip1_chr5_37146267_R | Primer3: not found at this Tm | N.d. | 0.05 |
| mm4_exon_Gm8425_chr16_36779053_F | TCGTCGGCAGCGTCTCAAATGGGCCCGCTACATT | 60.0 | 0.05 |
| mm4_exon_Gm8425_chr16_36779053_R | GTCTCGTGGGCTCGGCTCTGGCCTGTACTTGTGGG | 60.0 | 0.05 |
| mm4_intergenic_Gm9144|Apob_chr12_7926841_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm9144|Apob_chr12_7926841_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Cngb3|Gm11877_chr4_19293534_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Cngb3|Gm11877_chr4_19293534_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Olfr835|Olfr836_chr9_19064736_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Olfr835|Olfr836_chr9_19064736_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm22031|Gm4823_chr15_3661951_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm22031|Gm4823_chr15_3661951_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm9959|Gm9386_chr17_80749718_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm9959|Gm9386_chr17_80749718_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Pfas|Gm25819_chr11_68995048_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Pfas|Gm25819_chr11_68995048_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_1700022A21Rik|Nub1_chr5_24654423_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_1700022A21Rik|Nub1_chr5_24654423_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Pacsin2|Ttll1_chr15_83475318_F | TCGTCGGCAGCGTCGACCCCAGAGTCTTGAAAGCA | 59.9 | 0.04 |
| mm4_intergenic_Pacsin2|Ttll1_chr15_83475318_R | GTCTCGTGGGCTCGGACAAGGCTCATTCACTGCTCA | 59.9 | 0.04 |
| mm4_intergenic_Gm12057|RP23-242C19.7/B3gnt2_chr11_22820811_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm12057|RP23-242C19.7/B3gnt2_chr11_22820811_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intron_Cers6_chr2_68905126_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intron_Cers6_chr2_68905126_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_exon_Scube3_chr17_28169120_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_exon_Scube3_chr17_28169120_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Rpl31-ps15|Gm26404_chr3_37758367_F | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_intergenic_Rpl31-ps15|Gm26404_chr3_37758367_R | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_intergenic_Gm15230|Gm15231_chrX_166883068_F | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_intergenic_Gm15230|Gm15231_chrX_166883068_R | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_exon_Cep89_chr7_35397110_F | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_exon_Cep89_chr7_35397110_R | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_intergenic_Pax2|Fam178a_chr19_44847017_F | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intergenic_Pax2|Fam178a_chr19_44847017_R | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intron_Pde4a_chr9_21176984_F | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intron_Pde4a_chr9_21176984_R | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intron_Rbfox1_chr16_6868322_F | TCGTCGGCAGCGTCCAGTTTGTTCTTTGCTGGGCA | 59.8 | 0.02 |
| mm4_intron_Rbfox1_chr16_6868322_R | GTCTCGTGGGCTCGGTGATCAAATGGCAGCAACAGA | 58.4 | 0.02 |
| mm4_intron_Grip1_chr10_120039936_F | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intron_Grip1_chr10_120039936_R | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intergenic_Znhit6|Cyr61_chr3_145613937_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Znhit6|Cyr61_chr3_145613937_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Gm11676|Gm11677_chr11_111719708_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Gm11676|Gm11677_chr11_111719708_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intron_Mef2d_chr3_88148367_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intron_Mef2d_chr3_88148367_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Pabpc4|Bmp8a_chr4_123303117_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Pabpc4|Bmp8a_chr4_123303117_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_exon_Sdk1_chr5_142215344_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_exon_Sdk1_chr5_142215344_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Rybp|Gm23234_chr6_100407476_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Rybp|Gm23234_chr6_100407476_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm3_intergenic_Olfr43|Olfr404-ps1_chr11_74212498_F | TCGTCGGCAGCGTCTGGCTCTTCTATAGCTGTTCTTT | 57.3 | 0.00 |
| mm3_intergenic_Olfr43|Olfr404-ps1_chr11_74212498_R | GTCTCGTGGGCTCGGACACAGAAGAGCAACACCCT | 59.1 | 0.00 |
| mm4_intron_Hecw2_chr1_53983184_F | TCGTCGGCAGCGTCCTGAGTGTGAGCTGCTACCC | 60.1 | 0.00 |
| mm4_intron_Hecw2_chr1_53983184_R | GTCTCGTGGGCTCGGCCTCCAAACCTCGTCAGCAT | 60.0 | 0.00 |
| mm4_intergenic_Adtrp|Gm22157_chr13_41849288_F | Primer3: not found at this Tm | N.d. | 0.00 |
| mm4_intergenic_Adtrp|Gm22157_chr13_41849288_R | Primer3: not found at this Tm | N.d. | 0.00 |
| mm4_intergenic_Cpz|Plk-ps1_chr5_35542116_F | Primer3: not found at this Tm | N.d. | 0.00 |
| mm4_intergenic_Cpz|Plk-ps1_chr5_35542116_R | Primer3: not found at this Tm | N.d. | 0.00 |
| mm4_intergenic_Gm22567|Malt1_chr18_65430914_F | TCGTCGGCAGCGTCGAACTCTTCGCTCCTTGCCA | 60.3 | 0.00 |
| mm4_intergenic_Gm22567|Malt1_chr18_65430914_R | GTCTCGTGGGCTCGGCTGCGAAACAGGTTCCCC | 58.3 | 0.00 |
| mm4_intron_Anks1b_chr10_90702510_F | Primer3: not found at this Tm | N.d. | 0.00 |
| mm4_intron_Anks1b_chr10_90702510_R | Primer3: not found at this Tm | N.d. | 0.00 |
These only list off-targets that have primers in the table above. Primers underlined, off-targets in bold.
| mm4_intron_C2cd2_chr16_97909395 |
AGGCTTCGGTTGCTAATTCACCCTGTAGATGGACTAGTAGGAATGTACTTCACTCACCCACTCTCTGAGGCTACTTTTTT TTGGCAATATGAATATATTTAATGCTGAACTTCTGTAATGTCTGCATCTCTGTCCCTCCT |
| mm4_intron_Ntrk2_chr13_58955411 |
GCTTGTGTGTGCTGACATGGTGCTGAGTGACACCATGCACTCAGCCACATTCTTCCAGTCGATTGATTGACAGGCAGCCA CCACTAACTCTTGGATAAAGGGTTTCTCTGCATGGAAGGAGGGTGTGGCTATCAAGTGC |
| mm4_intron_Asb1_chr1_91542337 |
GTGAACCTTGTGCTGCCTTGGTCTCAGCCCAGTCAATGGACTTTCAGGCCGCAGAGGGCAAGGATACTTAGTCGTCACCA CCGTGTACCTATAGGAGAATCTGTAGGGGCTCAACGGA |
| mm4_intron_Lphn3_chr5_81321036 |
TGTTTCCTCACCAAAGCAACATCTTGTATATTAGTGTCTTTTTGTGCCACCTTGCACCTGTCGATGTACAAGCAAGCAAA GAGAGGGAATCCAGCAGAAAAACTGAGATTTGAACCTCAGTGTGGAAGGC |
| mm3_intergenic_Gm25148|Tifab_chr13_56171309 |
TAGATGGATGACCTGGGGCAGTGGAGAGTGACTACAACCCAGAGGCCGTGGGCAGGAGCAGGGGAAGCAGGAGCCAGTGG ATGGAGTAGGAGGCAGGTGCCCCAGCTGGTTTGG |
| mm4_intergenic_Adra2c|E130018O15Rik_chr5_35314570 | CCCGAGGGATGGAATGATGGCCATGCCCATGGATGGGCTAGCAGGCTGGATAGAGCCACTGCAGGGCTAAGAACA |
| mm4_intergenic_Irx1|Rpl9-ps4_chr13_72231918 |
TAAAAGCCCATCGTCCCCACTTCAGTGTTGGGATTCTGCCTACTAGACCATTGATAGGTGGCTTTGGAGACATGGTTCTA CCAACCCAGGAGGTTGGGAGTGTTGTAGGCATGCT |
| mm4_exon_Gm8425_chr16_36779053 |
TCAAATGGGCCCGCTACATTAGGCTCAAAATACCTCCTGCCATTAACCAGTTCACCCAGGCGCTGGACAAGCAGACAGCT ACTCAGCTGCTTAAGCTTGCCCACAAGTACAGGCCAGAG |
| mm4_intergenic_Pacsin2|Ttll1_chr15_83475318 | GACCCCAGAGTCTTGAAAGCAAAGACCTGGCTGCTTGTCCATAGCCAGGCTGAGCAGTGAATGAGCCTTGT |
| mm4_intron_Rbfox1_chr16_6868322 |
CAGTTTGTTCTTTGCTGGGCACCTTTCTCCTGTTTAGAGTGTCATCCAACACCTCTGTCTGCTAGTGCATTGAAGGGAAT TGTTTTCTGTTGCTGCCATTTGATCA |
| mm3_intergenic_Olfr43|Olfr404-ps1_chr11_74212498 |
TGGCTCTTCTATAGCTGTTCTTTATATAGATTTTGGTCTCAAGTCTGGTCCATGGACTATCAGACAGAAATATCATGGTA TCCTAATGATTAATGTCTTTTTTCTTGGTAAACACATTTAAGGGTGTTGCTCTTCTGTGT |
| mm4_intron_Hecw2_chr1_53983184 |
CTGAGTGTGAGCTGCTACCCAAAAGACCCAATCCATGGACTAACAGGAAGGTCACTCAGCTGAGCCCTGAATGATGCTGA CGAGGTTTGGAGG |
| mm4_intergenic_Gm22567|Malt1_chr18_65430914 |
GAACTCTTCGCTCCTTGCCAGTGGGGAAGCGCCACCCAAGCCCCGCCCATACACTAGCAGGCAGGACCCGCCCCCGGAGC CCGCCCCTCTTCTCCGCTGGCGCCCCGCGCGGCTTCACAGGGGGAACCTGTTTCGCAG |