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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_52314008_F | Primer3: not found at this Tm | N.d. | 1.00 |
| ontarget_mm0_exon_5730457N03Rik/Evx1_chr6_52314008_R | Primer3: not found at this Tm | N.d. | 1.00 |
| mm4_intergenic_4930564G21Rik|Grip1_chr10_119761855_F | Primer3: not found at this Tm | N.d. | 0.41 |
| mm4_intergenic_4930564G21Rik|Grip1_chr10_119761855_R | Primer3: not found at this Tm | N.d. | 0.41 |
| mm4_intron_Myo16_chr8_10439860_F | Primer3: not found at this Tm | N.d. | 0.39 |
| mm4_intron_Myo16_chr8_10439860_R | Primer3: not found at this Tm | N.d. | 0.39 |
| mm4_intergenic_Rnf214|Rnf214/Pcsk7_chr9_45906302_F | Primer3: not found at this Tm | N.d. | 0.38 |
| mm4_intergenic_Rnf214|Rnf214/Pcsk7_chr9_45906302_R | Primer3: not found at this Tm | N.d. | 0.38 |
| mm4_intron_Tmem241_chr18_12109978_F | Primer3: not found at this Tm | N.d. | 0.38 |
| mm4_intron_Tmem241_chr18_12109978_R | Primer3: not found at this Tm | N.d. | 0.38 |
| mm3_exon_Gm20403/Baz1a_chr12_54986310_F | Primer3: not found at this Tm | N.d. | 0.25 |
| mm3_exon_Gm20403/Baz1a_chr12_54986310_R | Primer3: not found at this Tm | N.d. | 0.25 |
| mm4_intergenic_Mpp5|Gm25225_chr12_78749153_F | Primer3: not found at this Tm | N.d. | 0.24 |
| mm4_intergenic_Mpp5|Gm25225_chr12_78749153_R | Primer3: not found at this Tm | N.d. | 0.24 |
| mm4_intergenic_Zfp516|Gm24720_chr18_83316789_F | Primer3: not found at this Tm | N.d. | 0.23 |
| mm4_intergenic_Zfp516|Gm24720_chr18_83316789_R | Primer3: not found at this Tm | N.d. | 0.23 |
| mm4_intron_Scaf8_chr17_3115624_F | TCGTCGGCAGCGTCAGGGCTGAGGGAAGGCAG | 61.0 | 0.22 |
| mm4_intron_Scaf8_chr17_3115624_R | GTCTCGTGGGCTCGGGGGTGGGAGGTGATAGTCCT | 60.0 | 0.22 |
| mm4_intergenic_4930419G24Rik|Pex5l_chr3_33122145_F | Primer3: not found at this Tm | N.d. | 0.21 |
| mm4_intergenic_4930419G24Rik|Pex5l_chr3_33122145_R | Primer3: not found at this Tm | N.d. | 0.21 |
| mm4_intron_Gm9994_chr1_93934074_F | Primer3: not found at this Tm | N.d. | 0.20 |
| mm4_intron_Gm9994_chr1_93934074_R | Primer3: not found at this Tm | N.d. | 0.20 |
| mm4_intron_Gm6086_chr1_94002351_F | TCGTCGGCAGCGTCGGACCAAGGCATCTCCTCTG | 59.8 | 0.20 |
| mm4_intron_Gm6086_chr1_94002351_R | GTCTCGTGGGCTCGGACCCTAGCTAGGATGGGACA | 59.0 | 0.20 |
| mm4_intron_Gal3st2_chr1_93867906_F | TCGTCGGCAGCGTCTCTACAGAGGGACCAAGGCA | 59.8 | 0.20 |
| mm4_intron_Gal3st2_chr1_93867906_R | GTCTCGTGGGCTCGGACCCTAGCTAGGATGGGACA | 59.0 | 0.20 |
| mm4_intergenic_Odf3b|Klhdc7b_chr15_89384909_F | Primer3: not found at this Tm | N.d. | 0.19 |
| mm4_intergenic_Odf3b|Klhdc7b_chr15_89384909_R | Primer3: not found at this Tm | N.d. | 0.19 |
| mm4_intron_Cacna1h_chr17_25408614_F | TCGTCGGCAGCGTCCTCCATCAGCAGCCACGTTA | 60.1 | 0.16 |
| mm4_intron_Cacna1h_chr17_25408614_R | GTCTCGTGGGCTCGGTGAAGCAACAGCCTTGGGAA | 60.1 | 0.16 |
| mm4_intron_Qdpr_chr5_45435941_F | TCGTCGGCAGCGTCGAGCTGGGTGTCTGGGAAAA | 59.8 | 0.15 |
| mm4_intron_Qdpr_chr5_45435941_R | GTCTCGTGGGCTCGGTGTGCTTAGCATGCCTTCGT | 60.3 | 0.15 |
| mm4_exon_Kif17_chr4_138262479_F | Primer3: not found at this Tm | N.d. | 0.15 |
| mm4_exon_Kif17_chr4_138262479_R | Primer3: not found at this Tm | N.d. | 0.15 |
| mm4_intergenic_Gm15401|Tcf7l1_chr6_72643370_F | Primer3: not found at this Tm | N.d. | 0.11 |
| mm4_intergenic_Gm15401|Tcf7l1_chr6_72643370_R | Primer3: not found at this Tm | N.d. | 0.11 |
| mm4_intron_Nkx6-3_chr8_23154497_F | TCGTCGGCAGCGTCGCAGAACCAGAGCGCTAAGA | 60.1 | 0.11 |
| mm4_intron_Nkx6-3_chr8_23154497_R | GTCTCGTGGGCTCGGCCCCTCCACTCCAGTTCTCA | 60.5 | 0.11 |
| mm4_exon_Znrf3_chr11_5278013_F | TCGTCGGCAGCGTCATGTGCAATCTGGACCCTGG | 60.0 | 0.11 |
| mm4_exon_Znrf3_chr11_5278013_R | GTCTCGTGGGCTCGGAGGAAGAGTTTGGCCGAGTG | 59.9 | 0.11 |
| mm4_intergenic_Nup214|Gm16534/Fam78a/Nup214_chr2_32051991_F | TCGTCGGCAGCGTCAGGTGTTGGGTTTAGGGCTG | 59.8 | 0.11 |
| mm4_intergenic_Nup214|Gm16534/Fam78a/Nup214_chr2_32051991_R | GTCTCGTGGGCTCGGCTGGAGCCTTCAGTAGCAGG | 59.8 | 0.11 |
| mm4_intergenic_Top2a|Igfbp4_chr11_99039389_F | TCGTCGGCAGCGTCTGGAGAGTCATGCTTGAGCC | 59.7 | 0.10 |
| mm4_intergenic_Top2a|Igfbp4_chr11_99039389_R | GTCTCGTGGGCTCGGCAACCCCAACACCTCCTCTG | 60.2 | 0.10 |
| mm4_intron_BC034090_chr1_155226593_F | TCGTCGGCAGCGTCACATTCTCCCATGGTGCCTG | 60.0 | 0.09 |
| mm4_intron_BC034090_chr1_155226593_R | GTCTCGTGGGCTCGGGATGTGCTCATCCTGCAGGG | 60.8 | 0.09 |
| mm4_intergenic_Gm23534|Gm22727_chr1_137671261_F | TCGTCGGCAGCGTCTGGATGGGTACGCATTGCTT | 60.0 | 0.07 |
| mm4_intergenic_Gm23534|Gm22727_chr1_137671261_R | GTCTCGTGGGCTCGGTCCCCAGAGAACGAACTGAA | 58.2 | 0.07 |
| mm4_intergenic_Mageb4|Gm8839_chrX_86266355_F | TCGTCGGCAGCGTCTGTGGATATTGGGGAAGCGG | 59.8 | 0.07 |
| mm4_intergenic_Mageb4|Gm8839_chrX_86266355_R | GTCTCGTGGGCTCGGCTGCCAGTCTGCACCTCTAC | 60.1 | 0.07 |
| mm3_intergenic_Ubr5|Odf1_chr15_38178303_F | Primer3: not found at this Tm | N.d. | 0.07 |
| mm3_intergenic_Ubr5|Odf1_chr15_38178303_R | Primer3: not found at this Tm | N.d. | 0.07 |
| mm4_intergenic_Gm15486|Sec16b_chr1_157518386_F | TCGTCGGCAGCGTCCCAGAGGCTCTTTCCAGCTC | 60.1 | 0.07 |
| mm4_intergenic_Gm15486|Sec16b_chr1_157518386_R | GTCTCGTGGGCTCGGCAGACACATATTCTGCACCCC | 58.7 | 0.07 |
| mm4_intergenic_Hoxc4|D930007P13Rik_chr15_103102479_F | TCGTCGGCAGCGTCATCCTTCCTTTCAGCTGCCC | 60.0 | 0.06 |
| mm4_intergenic_Hoxc4|D930007P13Rik_chr15_103102479_R | GTCTCGTGGGCTCGGCGATCCACAGAGCATCCGAG | 60.3 | 0.06 |
| mm4_intergenic_Cpsf6|Gm9002_chr10_117381753_F | Primer3: not found at this Tm | N.d. | 0.06 |
| mm4_intergenic_Cpsf6|Gm9002_chr10_117381753_R | Primer3: not found at this Tm | N.d. | 0.06 |
| mm4_intergenic_Igdcc3|Gm25313_chr9_65188735_F | Primer3: not found at this Tm | N.d. | 0.05 |
| mm4_intergenic_Igdcc3|Gm25313_chr9_65188735_R | Primer3: not found at this Tm | N.d. | 0.05 |
| mm4_intron_Fank1_chr7_133853427_F | TCGTCGGCAGCGTCGTTTCCCAGGTCACCGAGAC | 60.3 | 0.05 |
| mm4_intron_Fank1_chr7_133853427_R | GTCTCGTGGGCTCGGAAGAGTCCATCGGTTCAGCC | 59.7 | 0.05 |
| mm4_intergenic_Mir2136|Gm25904_chr9_104473197_F | TCGTCGGCAGCGTCGCCTCACTTTTCTCTGCACC | 59.1 | 0.05 |
| mm4_intergenic_Mir2136|Gm25904_chr9_104473197_R | GTCTCGTGGGCTCGGACCTGACCCCTTGAGTCTCT | 59.5 | 0.05 |
| mm4_intron_Hsp90ab1_chr17_45572959_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intron_Hsp90ab1_chr17_45572959_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm26068|Atrnl1_chr19_58054740_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intergenic_Gm26068|Atrnl1_chr19_58054740_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intron_Plekha7_chr7_116148134_F | TCGTCGGCAGCGTCGCCACAATGCTGAAGAGAGC | 59.5 | 0.04 |
| mm4_intron_Plekha7_chr7_116148134_R | GTCTCGTGGGCTCGGTCCATTGCTTCTGGCTCAGG | 60.0 | 0.04 |
| mm4_intron_Atg2b_chr12_105643416_F | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_intron_Atg2b_chr12_105643416_R | Primer3: not found at this Tm | N.d. | 0.04 |
| mm4_exon_Bicd1_chr6_149409175_F | TCGTCGGCAGCGTCAGAACCGGAGTCTCTTGCTG | 59.3 | 0.03 |
| mm4_exon_Bicd1_chr6_149409175_R | GTCTCGTGGGCTCGGGCATGGAGACTGCAAGGTGA | 60.3 | 0.03 |
| mm3_exon_Chic2_chr5_75044633_F | TCGTCGGCAGCGTCCAACCACAGCAACAGCCC | 59.2 | 0.03 |
| mm3_exon_Chic2_chr5_75044633_R | GTCTCGTGGGCTCGGGAGACCGTCCGCGTGAAC | 60.8 | 0.03 |
| mm4_intron_Slc25a29_chr12_108832432_F | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_intron_Slc25a29_chr12_108832432_R | Primer3: not found at this Tm | N.d. | 0.03 |
| mm4_intergenic_Gm8935|Gm2464_chr3_13308306_F | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intergenic_Gm8935|Gm2464_chr3_13308306_R | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intergenic_Ighv1-26|Ighv1-30_chr12_114802619_F | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intergenic_Ighv1-26|Ighv1-30_chr12_114802619_R | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intron_Rbpms_chr8_33922250_F | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intron_Rbpms_chr8_33922250_R | Primer3: not found at this Tm | N.d. | 0.02 |
| mm4_intron_Gm3898_chr9_43832325_F | TCGTCGGCAGCGTCGGTGACTATGTGGGGAGCAC | 60.1 | 0.02 |
| mm4_intron_Gm3898_chr9_43832325_R | GTCTCGTGGGCTCGGAGCTCTAGGTGGAGACCCTG | 60.0 | 0.02 |
| mm4_intergenic_Gm1862|Gm14591_chrX_53884653_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Gm1862|Gm14591_chrX_53884653_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_exon_Atf1_chr15_100227799_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_exon_Atf1_chr15_100227799_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intron_Unc79_chr12_103165327_F | TCGTCGGCAGCGTCCGCTCATCTCACAGGCATCT | 59.8 | 0.01 |
| mm4_intron_Unc79_chr12_103165327_R | GTCTCGTGGGCTCGGAGTGACGCCTCCTTGCTTTT | 60.1 | 0.01 |
| mm4_intergenic_Bub1b|Pak6_chr2_118650765_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Bub1b|Pak6_chr2_118650765_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_exon_Fbxo8_chr8_56551182_F | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_exon_Fbxo8_chr8_56551182_R | Primer3: not found at this Tm | N.d. | 0.01 |
| mm4_intergenic_Saysd1|Kcnk5_chr14_20117372_F | TCGTCGGCAGCGTCTTCTCCACTCTGTTGCAGGC | 60.2 | 0.00 |
| mm4_intergenic_Saysd1|Kcnk5_chr14_20117372_R | GTCTCGTGGGCTCGGAATGCAAGCAGGTCTCCCAA | 59.8 | 0.00 |
| mm4_intergenic_Gm17231|Arid3b_chr9_57780628_F | Primer3: not found at this Tm | N.d. | 0.00 |
| mm4_intergenic_Gm17231|Arid3b_chr9_57780628_R | Primer3: not found at this Tm | N.d. | 0.00 |
| mm4_exon_1700024N20Rik/Prosc_chr8_27042868_F | TCGTCGGCAGCGTCGGATCGCTGGGTTTCCATGG | 61.1 | 0.00 |
| mm4_exon_1700024N20Rik/Prosc_chr8_27042868_R | GTCTCGTGGGCTCGGGCAATGACGAATGGCGAGAC | 59.9 | 0.00 |
These only list off-targets that have primers in the table above. Primers underlined, off-targets in bold.
| mm4_intron_Scaf8_chr17_3115624 |
AGGGCTGAGGGAAGGCAGGGGGAGGGCCGGTCCCCAAGGCGGCCGCGCCACGCTCCCGGGTGGGTTCCACCGAGGACTAT CACCTCCCACCC |
| mm4_intron_Gm6086_chr1_94002351 |
GGACCAAGGCATCTCCTCTGAGGAGCTCCTGAGCTCTTTGTCTATACAGACTTTCAGGGAGTGTGGTGGGGACATGACTG ACACACATCAGAGGTGACATGTCCCATCCTAGCTAGGGT |
| mm4_intron_Gal3st2_chr1_93867906 |
TCTACAGAGGGACCAAGGCATCTCCTCTGGGGAGCTCCTGAGCTCTTTGTCTATACAGACTTTCAGGGAGTGTGGTGGGG ACACGACTGACACACATCAGAGGTGACATGTCCCATCCTAGCTAGGGT |
| mm4_intron_Cacna1h_chr17_25408614 |
CTCCATCAGCAGCCACGTTATTAATAGTGGTGACATTTAAAAAAAGATGGTCCAGANNNNNNNNNNNNNNNNNNNNNNNN AAAATCTGATTTCAAAGTGAAGCCTGCTGGACCTTGGTTTCCCAAGGCTGTTGCTTCA |
| mm4_intron_Qdpr_chr5_45435941 |
GAGCTGGGTGTCTGGGAAAAACAAAAGTGACATTTGGAGACATTTAAGGGTTTCTATTTTCATCACAGAAGTCTGTCCTG CCTTTGCCTTCCCAGGAGTGTGGAGTGGATGAGAACGAAGGCATGCTAAGCACA |
| mm4_intron_Nkx6-3_chr8_23154497 |
GCAGAACCAGAGCGCTAAGATGACCTTGGCCCTGACTGCTAGGGANNNNNNNNNNNNNNNNNNNNNNTTACCCCACCCAT CCCTGAGACTCAGGTTTCTGAGAACTGGAGTGGAGGGG |
| mm4_exon_Znrf3_chr11_5278013 |
ATGTGCAATCTGGACCCTGGGATGACTTCCTGGGAGCCTGACTGGGAAGAAGGGTCATTTCTGTCTCTGGCAACCCAGGT GGTCCCCATTCTGTAACCGAGTTTTCAGACCCACTCGGCCAAACTCTTCCT |
| mm4_intergenic_Nup214|Gm16534/Fam78a/Nup214_chr2_32051991 |
AGGTGTTGGGTTTAGGGCTGCCGGCCTGCGGGAGGCTGAGTAAGAGATGATTGGCCTTCCCGGCAGCAGGGCGTGGAGAC ACTTGCCTCATTTCTGTGGTGGTGTCTCCAGCTGCCTGCTACTGAAGGCTCCAG |
| mm4_intergenic_Top2a|Igfbp4_chr11_99039389 |
TGGAGAGTCATGCTTGAGCCCTGACTTCCCGGGAATGGGGTGGGGGTGGGGAGACACCAGAGGAAGGTTTGGGGGCAGAG GAGGTGTTGGGGTTG |
| mm4_intron_BC034090_chr1_155226593 |
ACATTCTCCCATGGTGCCTGACGGTACCAACAATCCACAAGGTTACCCCCAGTCAGGCTCCCGGGAAGACACGTGACAAG CCCCAAGCCCTGCAGGATGAGCACATC |
| mm4_intergenic_Gm23534|Gm22727_chr1_137671261 |
TGGATGGGTACGCATTGCTTCTTGTCAACTGCTTCTGCTCCACCAGCTGTGTGGAGCACAGCTAACTTCCAGGGAGCTTG GCAAGGCAGGCATTCAGTTCGTTCTCTGGGGA |
| mm4_intergenic_Mageb4|Gm8839_chrX_86266355 |
TGTGGATATTGGGGAAGCGGTCTTACCTGTGTCTCTGGTTAGAGCAGACTTCCTGGGAGACTGGCAGGGCTGTGGGTACA GGTAGCAAGCATGCTGCTCTGCCACAGGTACAGGTAGAGGTGCAGACTGGCAG |
| mm4_intergenic_Gm15486|Sec16b_chr1_157518386 |
CCAGAGGCTCTTTCCAGCTCATTGGCTCTGAGTGCTTGAATGTCTGCTCTTGTTCAGAATATGTCTCTCAGACTTCCCCA CTGCGTGGCGAGGAATGACTCAGTGTGGGGGTGCAGAATATGTGTCTG |
| mm4_intergenic_Hoxc4|D930007P13Rik_chr15_103102479 |
ATCCTTCCTTTCAGCTGCCCGGTGTTCTCCACCCCTGCGTTAGCACTCTGTAGTCTGGGGACCTCCCGGGAACGTCCCGG GGCTAGCAACCAAGGGGGCTCTCGGATGCTCTGTGGATCG |
| mm4_intron_Fank1_chr7_133853427 | GTTTCCCAGGTCACCGAGACACCCCCCCCCCCACGCTCCCCTGAAGTTATGGGCTGAACCGATGGACTCTT |
| mm4_intergenic_Mir2136|Gm25904_chr9_104473197 |
GCCTCACTTTTCTCTGCACCTATTGGTTCTCATGAAGTTAGAATTTATGTAGAAACCTTACTGGCAGCAGTGACTGCCCT GGAGAGTGGAGGGGACTAGAAGAGACTCAAGGGGTCAGGT |
| mm4_intron_Plekha7_chr7_116148134 |
GCCACAATGCTGAAGAGAGCTTTGGGTTAAAGAAAATCAACCGAAATGACTTGCAGGAAGCGTGGGGCGGTCAGGAGAGC AACTGCTTAAGCAAGTGATGGTGCCGGCCCTGAGCCAGAAGCAATGGA |
| mm4_exon_Bicd1_chr6_149409175 |
AGAACCGGAGTCTCTTGCTGCGCGAGTTGCCCCTTGGCCATGCAGCTTGGCTGCTGACTGCCCCGGCGCGTGGCCCGGAG GATCCCGGCTGCGGTCATCACCTTGCAGTCTCCATGC |
| mm3_exon_Chic2_chr5_75044633 |
CAACCACAGCAACAGCCCGAGCCACCCGCAGCAGGAGCAGCCGGCTACCGCATCGCGAGACTTCCCGGGAGCNNNNNNNN NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNCGTTCACGCGGACGGTCTC |
| mm4_intron_Gm3898_chr9_43832325 |
GGTGACTATGTGGGGAGCACCCCCACCAAGCTCCAGGGCAGTCATCTGGAGCAGGGCGTGGGGTGGTCTGTACAGCTCCC TCTTTCTTTTCCAGGGTCTCCACCTAGAGCT |
| mm4_intron_Unc79_chr12_103165327 |
CGCTCATCTCACAGGCATCTGAGACTTCGAGACTTCCCTGGAGCCTGGGATGGATCAGTTAGGTCTGACTCTTGTGCTTA TACCGGTTTAAAAGCAAGGAGGCGTCACT |
| mm4_intergenic_Saysd1|Kcnk5_chr14_20117372 |
TTCTCCACTCTGTTGCAGGCGAGCCTGTCAAAGAAGAGAACTGGGTGCTGATGATAGTAGAGCCACCCCACCTTCCTGGG AAGTCAATGGTGTGGGTCTGGCTTTGCTTGGGAGACCTGCTTGCATT |
| mm4_exon_1700024N20Rik/Prosc_chr8_27042868 |
GGATCGCTGGGTTTCCATGGCGGTCGGTCTGGCCTTGAGGATCGGGACTTGCCGGGAGGTTGGCTTGGTTGGAGAAAGGT GTTCATTTAGGGCAAGTGACTCGTATCCAGGCGTCTCGCCATTCGTCATTGC |