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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 |
---|---|---|---|
mm0_exon_Pop5_chr5_115238035_F | Primer3: not found at this Tm | N.d. | 1.00 |
mm0_exon_Pop5_chr5_115238035_R | Primer3: not found at this Tm | N.d. | 1.00 |
mm4_intergenic_Sec11a|Zfp592_chr7_80981225_F | Primer3: not found at this Tm | N.d. | 0.53 |
mm4_intergenic_Sec11a|Zfp592_chr7_80981225_R | Primer3: not found at this Tm | N.d. | 0.53 |
mm4_intergenic_Gm12726|Gm12728_chr4_105710801_F | Primer3: not found at this Tm | N.d. | 0.36 |
mm4_intergenic_Gm12726|Gm12728_chr4_105710801_R | Primer3: not found at this Tm | N.d. | 0.36 |
mm4_intergenic_Rpl31-ps8|Tcea1-ps1_chr15_90813139_F | Primer3: not found at this Tm | N.d. | 0.36 |
mm4_intergenic_Rpl31-ps8|Tcea1-ps1_chr15_90813139_R | Primer3: not found at this Tm | N.d. | 0.36 |
mm4_exon_Fiz1_chr7_5008352_F | Primer3: not found at this Tm | N.d. | 0.32 |
mm4_exon_Fiz1_chr7_5008352_R | Primer3: not found at this Tm | N.d. | 0.32 |
mm4_exon_Nop56_chr2_130274459_F | TCGTCGGCAGCGTCTGGTCAAACCGGAGAAGGTG | 59.8 | 0.31 |
mm4_exon_Nop56_chr2_130274459_R | GTCTCGTGGGCTCGGTACGCAAATACCCCACCACC | 60.0 | 0.31 |
mm4_intergenic_Gm25905|Dcp1b_chr6_118989106_F | TCGTCGGCAGCGTCCACCAGGAGAGCAACCAGAG | 60.0 | 0.30 |
mm4_intergenic_Gm25905|Dcp1b_chr6_118989106_R | GTCTCGTGGGCTCGGAGGCAAAACTGACTCGGAGG | 59.9 | 0.30 |
mm4_exon_Ogg1_chr6_113331935_F | Primer3: not found at this Tm | N.d. | 0.24 |
mm4_exon_Ogg1_chr6_113331935_R | Primer3: not found at this Tm | N.d. | 0.24 |
mm4_intergenic_Nhsl2|Gm25828_chrX_101864982_F | Primer3: not found at this Tm | N.d. | 0.23 |
mm4_intergenic_Nhsl2|Gm25828_chrX_101864982_R | Primer3: not found at this Tm | N.d. | 0.23 |
mm4_exon_Slc43a2_chr11_75532621_F | TCGTCGGCAGCGTCGGTGTCTGCAGAACCGGAG | 60.3 | 0.20 |
mm4_exon_Slc43a2_chr11_75532621_R | GTCTCGTGGGCTCGGGGTTTTCCAACACAGCGGTG | 60.2 | 0.20 |
mm4_intron_Nrip1_chr16_76372649_F | Primer3: not found at this Tm | N.d. | 0.20 |
mm4_intron_Nrip1_chr16_76372649_R | Primer3: not found at this Tm | N.d. | 0.20 |
mm4_intergenic_Rd3/Gm26670|Traf5_chr1_191991591_F | Primer3: not found at this Tm | N.d. | 0.16 |
mm4_intergenic_Rd3/Gm26670|Traf5_chr1_191991591_R | Primer3: not found at this Tm | N.d. | 0.16 |
mm4_intergenic_Aim1/Gm10812|Speer5-ps1_chr10_44166802_F | Primer3: not found at this Tm | N.d. | 0.15 |
mm4_intergenic_Aim1/Gm10812|Speer5-ps1_chr10_44166802_R | Primer3: not found at this Tm | N.d. | 0.15 |
mm4_intergenic_Nat3|Psd3_chr8_67593348_F | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intergenic_Nat3|Psd3_chr8_67593348_R | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intron_Gm960_chr19_4631687_F | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intron_Gm960_chr19_4631687_R | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intron_Vac14_chr8_110686960_F | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intron_Vac14_chr8_110686960_R | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intron_6030443J06Rik_chr5_22552874_F | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intron_6030443J06Rik_chr5_22552874_R | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intron_Ror2_chr13_53121182_F | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intron_Ror2_chr13_53121182_R | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intron_4933426M11Rik_chr12_80793003_F | Primer3: not found at this Tm | N.d. | 0.11 |
mm4_intron_4933426M11Rik_chr12_80793003_R | Primer3: not found at this Tm | N.d. | 0.11 |
mm4_exon_C3_chr17_57224326_F | TCGTCGGCAGCGTCAGTGCAGGATGACAGTGACG | 60.0 | 0.11 |
mm4_exon_C3_chr17_57224326_R | GTCTCGTGGGCTCGGAGATTGAGGATGGTGTGGGG | 59.0 | 0.11 |
mm4_exon_Mia3_chr1_183369513_F | Primer3: not found at this Tm | N.d. | 0.10 |
mm4_exon_Mia3_chr1_183369513_R | Primer3: not found at this Tm | N.d. | 0.10 |
mm4_exon_Suds3_chr5_117115953_F | TCGTCGGCAGCGTCGACTCCCCTCGGCTCCGA | 62.8 | 0.09 |
mm4_exon_Suds3_chr5_117115953_R | GTCTCGTGGGCTCGGCTGATCAGGCTGTCACCGAG | 60.1 | 0.09 |
mm4_exon_Bax_chr7_45466869_F | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_exon_Bax_chr7_45466869_R | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_exon_0610010F05Rik_chr11_23633578_F | Primer3: not found at this Tm | N.d. | 0.08 |
mm4_exon_0610010F05Rik_chr11_23633578_R | Primer3: not found at this Tm | N.d. | 0.08 |
mm4_intergenic_Gm26703|Dspp_chr5_104156812_F | Primer3: not found at this Tm | N.d. | 0.08 |
mm4_intergenic_Gm26703|Dspp_chr5_104156812_R | Primer3: not found at this Tm | N.d. | 0.08 |
mm4_intergenic_Bicc1|Tfam_chr10_71167910_F | Primer3: not found at this Tm | N.d. | 0.06 |
mm4_intergenic_Bicc1|Tfam_chr10_71167910_R | Primer3: not found at this Tm | N.d. | 0.06 |
mm4_intergenic_Csrp1|Phlda3_chr1_135752264_F | TCGTCGGCAGCGTCCTGTAGGCATGGGCTGTTGA | 60.0 | 0.06 |
mm4_intergenic_Csrp1|Phlda3_chr1_135752264_R | GTCTCGTGGGCTCGGTGCTGAGATTGTTCTGGTCTGA | 59.3 | 0.06 |
mm4_intergenic_Pds5b|Kl_chr5_150864378_F | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_intergenic_Pds5b|Kl_chr5_150864378_R | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_exon_Gm19557/Obfc1_chr19_47515307_F | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_exon_Gm19557/Obfc1_chr19_47515307_R | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_intron_Galnt7_chr8_57652304_F | TCGTCGGCAGCGTCGACCACAGGAGAGGCTGGA | 60.6 | 0.05 |
mm4_intron_Galnt7_chr8_57652304_R | GTCTCGTGGGCTCGGGTGCAGGCCTGAAGGAGG | 60.0 | 0.05 |
mm4_intron_Hmgb3_chrX_71557164_F | TCGTCGGCAGCGTCTTATGGGTGTCCGGAGCAAC | 60.0 | 0.05 |
mm4_intron_Hmgb3_chrX_71557164_R | GTCTCGTGGGCTCGGCTGAATTAAAGCCCAGCCGC | 59.8 | 0.05 |
mm3_intergenic_Rragc|Gm12916_chr4_124240638_F | Primer3: not found at this Tm | N.d. | 0.05 |
mm3_intergenic_Rragc|Gm12916_chr4_124240638_R | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_intergenic_Robo3|Msantd2_chr9_37460539_F | TCGTCGGCAGCGTCGGCCAAAGTAATGAGCCGCT | 61.0 | 0.04 |
mm4_intergenic_Robo3|Msantd2_chr9_37460539_R | GTCTCGTGGGCTCGGGAGCAGGTGTGAAGCGAGG | 60.7 | 0.04 |
mm4_intron_Bicd2_chr13_49382526_F | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intron_Bicd2_chr13_49382526_R | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intergenic_Guk1|2610507I01Rik_chr11_59197273_F | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intergenic_Guk1|2610507I01Rik_chr11_59197273_R | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intergenic_Gm5449|Drd1a_chr13_53779838_F | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intergenic_Gm5449|Drd1a_chr13_53779838_R | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_exon_Zfp12_chr5_143235362_F | Primer3: not found at this Tm | N.d. | 0.02 |
mm4_exon_Zfp12_chr5_143235362_R | Primer3: not found at this Tm | N.d. | 0.02 |
mm4_exon_Hcn1_chr13_117602765_F | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_exon_Hcn1_chr13_117602765_R | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intergenic_Gm24073|Mir17hg_chr14_115042223_F | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intergenic_Gm24073|Mir17hg_chr14_115042223_R | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intergenic_Gm26766|Gm16136_chr15_37120841_F | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Gm26766|Gm16136_chr15_37120841_R | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intron_Ephb2_chr4_136835231_F | TCGTCGGCAGCGTCAAGAGTGATGCCCATCGCC | 60.1 | 0.00 |
mm4_intron_Ephb2_chr4_136835231_R | GTCTCGTGGGCTCGGCCGTGGAGTGGAGCGAAG | 60.1 | 0.00 |
mm4_exon_Ina/Nt5c2_chr19_47014691_F | TCGTCGGCAGCGTCCAACCCCGCGCCTTAAAAG | 59.7 | 0.00 |
mm4_exon_Ina/Nt5c2_chr19_47014691_R | GTCTCGTGGGCTCGGATAGTGCTCCGATCCGAAGC | 59.6 | 0.00 |
These only list off-targets that have primers in the table above. Primers underlined, off-targets in bold.
mm4_exon_Nop56_chr2_130274459 |
TGGTCAAACCGGAGAAGGTGTGGTCTCCGCAGTCTGGTTCCGCGCGCCAGAGCGCGCTAGCCGCATTGCGAGCTGGGCAC GGAGCAGGCGCCATGGTGAGGAGTGGTGGTGGGGTATTTGCGTA |
mm4_intergenic_Gm25905|Dcp1b_chr6_118989106 |
CACCAGGAGAGCAACCAGAGCACCAAGTGATTGGCCTCACCATAGCGTCCAGGCTGGCAGATGCAAAACTGAAGCAGCAG AGACAGAGGTGTGCACTGGAGCGCTGGCCCCTCCGAGTCAGTTTTGCCT |
mm4_exon_Slc43a2_chr11_75532621 |
GGTGTCTGCAGAACCGGAGCCCGACCACCGCCACACCGCACCATGGCGCCCACCCTGGCCACTGCCCATCGGCGCCGCTG GTGGATGGCCTGCACCGCTGTGTTGGAAAACC |
mm4_exon_C3_chr17_57224326 |
AGTGCAGGATGACAGTGACGGAGACATACAGGGACTTCCCCACCAGGGCGTCGGCGTTGGAAGGCCGTACCCCCTCCATC AGCACCTTCCGGGTCAGCACTGCATCCCCCACACCATCCTCAATCT |
mm4_exon_Suds3_chr5_117115953 |
GACTCCCCTCGGCTCCGAGCTCCGCCCCCCACCCCGCACCCCGGCGCCCGCGCTGGGCAACGGGGGCCTCGCGCAGCCGG CTACTTCCGGGTCGTGGGCTGCTCGGTGACAGCCTGATCAG |
mm4_intergenic_Csrp1|Phlda3_chr1_135752264 |
CTGTAGGCATGGGCTGTTGAAGCTGATTTATCATCTGATCAATAAAGCAATTTAGAACTGAACTCTGCTTTCCTGTCTGT CTATGGGGGCCCGGCCACACCCTGGAGTCAGCGCTGGATCAGACCAGAACAATCTCAGCA |
mm4_intron_Galnt7_chr8_57652304 |
GACCACAGGAGAGGCTGGACCGCGGGGCCTGCTCCACCGGACCACGGCGGCGGCGCCGGCATCTCGCCTCCTTCAGGCCT GCAC |
mm4_intron_Hmgb3_chrX_71557164 |
TTATGGGTGTCCGGAGCAACGGGGACGTGGCCCGACACGGGCTGTCTGCTGCCGGCCTGGACGCCCGGGTGCGGGGGCCC GCCCCACCCGGGATACCGCGGCTGGGCTTTAATTCAG |
mm4_intergenic_Robo3|Msantd2_chr9_37460539 |
GGCCAAAGTAATGAGCCGCTAGGCAACTGGCCCCACGCCATTGTGTGCGCGCCGGGGCGAGGCCGGGTCTCCGCGGCGCG GTGGAGGGTGACCACGCGCGCCGCAGCGCCCCTCGCTTCACACCTGCTC |
mm4_intron_Ephb2_chr4_136835231 |
AAGAGTGATGCCCATCGCCCCATCCCCCGCTCAGTCCTGCTGGGGAGGGGACCCCATTGTACGCCGCGAGCGACCGGGCG GGCACGGACGCGAGGGTGGGGGACACGCGCGCTTCGCTCCACTCCACGG |
mm4_exon_Ina/Nt5c2_chr19_47014691 |
CAACCCCGCGCCTTAAAAGCCCCGCACACCGCCCCGCCGCACCCAGCCTTGCCGCACCTTCGCGTCCTCGCCAGGTCCCG CCGCAGCCGCGCACCCGGCCCCGACCCCGGCACCATGAGCTTCGGATCGGAGCACTAT |