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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_52314171_F | Primer3: not found at this Tm | N.d. | 1.00 |
ontarget_mm0_exon_5730457N03Rik/Evx1_chr6_52314171_R | Primer3: not found at this Tm | N.d. | 1.00 |
mm3_intron_Zbtb7c_chr18_75994536_F | Primer3: not found at this Tm | N.d. | 0.40 |
mm3_intron_Zbtb7c_chr18_75994536_R | Primer3: not found at this Tm | N.d. | 0.40 |
mm4_intron_Cdk14_chr5_5388180_F | Primer3: not found at this Tm | N.d. | 0.40 |
mm4_intron_Cdk14_chr5_5388180_R | Primer3: not found at this Tm | N.d. | 0.40 |
mm3_intergenic_5S_rRNA|Cops8_chr1_90587821_F | Primer3: not found at this Tm | N.d. | 0.33 |
mm3_intergenic_5S_rRNA|Cops8_chr1_90587821_R | Primer3: not found at this Tm | N.d. | 0.33 |
mm4_intron_Med16_chr10_79907907_F | TCGTCGGCAGCGTCCACTTCAGTCTGGGCCTCAG | 60.0 | 0.33 |
mm4_intron_Med16_chr10_79907907_R | GTCTCGTGGGCTCGGAGTATTGTGTGGCATGGGCT | 59.6 | 0.33 |
mm4_intergenic_Dnah5|Mir3964_chr15_28975090_F | Primer3: not found at this Tm | N.d. | 0.32 |
mm4_intergenic_Dnah5|Mir3964_chr15_28975090_R | Primer3: not found at this Tm | N.d. | 0.32 |
mm4_intron_Mettl15_chr2_109227338_F | Primer3: not found at this Tm | N.d. | 0.31 |
mm4_intron_Mettl15_chr2_109227338_R | Primer3: not found at this Tm | N.d. | 0.31 |
mm3_intergenic_Galnt13|Gm25489_chr2_54971077_F | Primer3: not found at this Tm | N.d. | 0.26 |
mm3_intergenic_Galnt13|Gm25489_chr2_54971077_R | Primer3: not found at this Tm | N.d. | 0.26 |
mm4_exon_Vdr_chr15_97859430_F | Primer3: not found at this Tm | N.d. | 0.23 |
mm4_exon_Vdr_chr15_97859430_R | Primer3: not found at this Tm | N.d. | 0.23 |
mm4_intron_Akr1c20_chr13_4501162_F | Primer3: not found at this Tm | N.d. | 0.23 |
mm4_intron_Akr1c20_chr13_4501162_R | Primer3: not found at this Tm | N.d. | 0.23 |
mm4_exon_Palld_chr8_61720535_F | TCGTCGGCAGCGTCACCATTCACACTTTGCCCCT | 59.8 | 0.19 |
mm4_exon_Palld_chr8_61720535_R | GTCTCGTGGGCTCGGTGGTCATTGCTGAGGCCTTT | 59.8 | 0.19 |
mm4_intron_Xylt1_chr7_117496014_F | Primer3: not found at this Tm | N.d. | 0.18 |
mm4_intron_Xylt1_chr7_117496014_R | Primer3: not found at this Tm | N.d. | 0.18 |
mm4_intergenic_Rasa1|Cox7c_chr13_85413950_F | Primer3: not found at this Tm | N.d. | 0.17 |
mm4_intergenic_Rasa1|Cox7c_chr13_85413950_R | Primer3: not found at this Tm | N.d. | 0.17 |
mm4_intergenic_Gm4823|Gm23228_chr15_3748714_F | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intergenic_Gm4823|Gm23228_chr15_3748714_R | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intron_Atp10d_chr5_72224585_F | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intron_Atp10d_chr5_72224585_R | Primer3: not found at this Tm | N.d. | 0.14 |
mm4_intergenic_Gm24436|Bre_chr5_31870840_F | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intergenic_Gm24436|Bre_chr5_31870840_R | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intergenic_Gm22825|Zfhx3_chr8_108111187_F | Primer3: not found at this Tm | N.d. | 0.12 |
mm4_intergenic_Gm22825|Zfhx3_chr8_108111187_R | Primer3: not found at this Tm | N.d. | 0.12 |
mm4_intergenic_Actb|Fscn1_chr5_142936443_F | TCGTCGGCAGCGTCGGACAATTACAGGCTGGGCT | 60.0 | 0.12 |
mm4_intergenic_Actb|Fscn1_chr5_142936443_R | GTCTCGTGGGCTCGGACATAGCAGCCTATCCCGGA | 60.1 | 0.12 |
mm4_intron_Nrg2_chr18_36118511_F | TCGTCGGCAGCGTCCCTCACAGAGACCTCCAGGA | 59.9 | 0.10 |
mm4_intron_Nrg2_chr18_36118511_R | GTCTCGTGGGCTCGGGCCTAGGGAAAAGGGCATGT | 60.0 | 0.10 |
mm4_intron_Cdc14a_chr3_116307187_F | TCGTCGGCAGCGTCACTAGCACAGTAGCTTCCAGC | 59.7 | 0.09 |
mm4_intron_Cdc14a_chr3_116307187_R | GTCTCGTGGGCTCGGTGTCTGATCCCAAACTCATGCT | 59.6 | 0.09 |
mm4_intron_Zmym4_chr4_126922543_F | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_intron_Zmym4_chr4_126922543_R | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_intergenic_Gm22848|Rp1_chr1_4025600_F | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_intergenic_Gm22848|Rp1_chr1_4025600_R | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_intergenic_Dancr/Snora26|Rasl11b_chr5_74097935_F | TCGTCGGCAGCGTCGCTCGCTTTTCACACCAAGG | 60.0 | 0.09 |
mm4_intergenic_Dancr/Snora26|Rasl11b_chr5_74097935_R | GTCTCGTGGGCTCGGTGTCCGAGAACCCTTGATGC | 60.0 | 0.09 |
mm4_intergenic_Gm5205|Sh2d4b_chr14_40693943_F | TCGTCGGCAGCGTCGGTAGTCAGGCTTCAGGCAT | 59.4 | 0.08 |
mm4_intergenic_Gm5205|Sh2d4b_chr14_40693943_R | GTCTCGTGGGCTCGGTGTCTTCAGGTGACTTTGCCT | 59.5 | 0.08 |
mm3_intergenic_Gm8298|C130079G13Rik_chr3_59913061_F | Primer3: not found at this Tm | N.d. | 0.08 |
mm3_intergenic_Gm8298|C130079G13Rik_chr3_59913061_R | Primer3: not found at this Tm | N.d. | 0.08 |
mm4_intron_Tex22_chr12_113083624_F | Primer3: not found at this Tm | N.d. | 0.06 |
mm4_intron_Tex22_chr12_113083624_R | Primer3: not found at this Tm | N.d. | 0.06 |
mm3_intergenic_Panx3|Gm8343_chr9_37679728_F | Primer3: not found at this Tm | N.d. | 0.06 |
mm3_intergenic_Panx3|Gm8343_chr9_37679728_R | Primer3: not found at this Tm | N.d. | 0.06 |
mm4_intron_Pitpnc1_chr11_107249066_F | TCGTCGGCAGCGTCCTGTTTCGCTGGCACTGATG | 59.8 | 0.06 |
mm4_intron_Pitpnc1_chr11_107249066_R | GTCTCGTGGGCTCGGTGCACACACACACAATTGGG | 59.5 | 0.06 |
mm4_intergenic_Mrps28|Gm15467_chr3_8850429_F | TCGTCGGCAGCGTCGGATAGCCTGCAGAGACGAC | 59.9 | 0.06 |
mm4_intergenic_Mrps28|Gm15467_chr3_8850429_R | GTCTCGTGGGCTCGGGACCCACCCTCTGATGAAGC | 60.1 | 0.06 |
mm4_intergenic_Rpl27a-ps2|Slitrk1_chr14_108819867_F | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_intergenic_Rpl27a-ps2|Slitrk1_chr14_108819867_R | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_intron_Scel_chr14_103590622_F | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_intron_Scel_chr14_103590622_R | Primer3: not found at this Tm | N.d. | 0.05 |
mm3_exon_Dixdc1_chr9_50698817_F | Primer3: not found at this Tm | N.d. | 0.04 |
mm3_exon_Dixdc1_chr9_50698817_R | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intergenic_Hipk3|Gm13870_chr2_104508381_F | TCGTCGGCAGCGTCTCACTGCTTGCCACTTCTGA | 59.5 | 0.04 |
mm4_intergenic_Hipk3|Gm13870_chr2_104508381_R | GTCTCGTGGGCTCGGCAGCCGACCCTTCTCCTTTT | 59.9 | 0.04 |
mm4_intron_Cacna1d_chr14_30271304_F | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intron_Cacna1d_chr14_30271304_R | Primer3: not found at this Tm | N.d. | 0.04 |
mm4_intergenic_Stox2|4930448N21Rik/Gm10083_chr8_47366683_F | Primer3: not found at this Tm | N.d. | 0.03 |
mm4_intergenic_Stox2|4930448N21Rik/Gm10083_chr8_47366683_R | Primer3: not found at this Tm | N.d. | 0.03 |
mm4_intron_Nos1ap_chr1_170423646_F | TCGTCGGCAGCGTCCCCAGAAGGACTTAGCAGAGC | 60.1 | 0.03 |
mm4_intron_Nos1ap_chr1_170423646_R | GTCTCGTGGGCTCGGTGGAGGCTGGAAAGGAGGTA | 59.8 | 0.03 |
mm4_intron_Slc9a9_chr9_95193065_F | TCGTCGGCAGCGTCGGAAGGCATGTCCACACAGA | 59.9 | 0.03 |
mm4_intron_Slc9a9_chr9_95193065_R | GTCTCGTGGGCTCGGCCTAGAGATGGCCTTGGGGA | 60.4 | 0.03 |
mm4_intergenic_Gm23653|Nnmt_chr9_48580114_F | TCGTCGGCAGCGTCTGTCAGAGATTGTCCATGTTGT | 57.6 | 0.03 |
mm4_intergenic_Gm23653|Nnmt_chr9_48580114_R | GTCTCGTGGGCTCGGAGCTGAGTCTGCTTTTAGTCTCT | 59.1 | 0.03 |
mm4_intergenic_Gm24254|4933436C20Rik_chr8_92201343_F | Primer3: not found at this Tm | N.d. | 0.03 |
mm4_intergenic_Gm24254|4933436C20Rik_chr8_92201343_R | Primer3: not found at this Tm | N.d. | 0.03 |
mm4_intergenic_Gm5973|Gm9915_chr1_42186377_F | TCGTCGGCAGCGTCATGGACTCACCTGGACCTGA | 59.8 | 0.03 |
mm4_intergenic_Gm5973|Gm9915_chr1_42186377_R | GTCTCGTGGGCTCGGTGCTCCAGGGTTGAAATGGT | 59.5 | 0.03 |
mm4_intergenic_Rps6-ps2|mmu-mir-8110_chr8_88909832_F | Primer3: not found at this Tm | N.d. | 0.02 |
mm4_intergenic_Rps6-ps2|mmu-mir-8110_chr8_88909832_R | Primer3: not found at this Tm | N.d. | 0.02 |
mm4_intergenic_Fto|Irx3_chr8_91711866_F | TCGTCGGCAGCGTCCTCCTGATTCCTGCTGCGAT | 59.8 | 0.02 |
mm4_intergenic_Fto|Irx3_chr8_91711866_R | GTCTCGTGGGCTCGGATCGGGCAGAGATTCCGTTC | 59.8 | 0.02 |
mm4_intergenic_Gm24069|Gm24434_chr12_39521785_F | TCGTCGGCAGCGTCGCTGGTGCTGAGAATATAGTGGT | 60.1 | 0.02 |
mm4_intergenic_Gm24069|Gm24434_chr12_39521785_R | GTCTCGTGGGCTCGGTGACTCAAAATACAACTTTCTCACA | 57.1 | 0.02 |
mm4_intergenic_Ntrk2|Agtpbp1_chr13_59360879_F | Primer3: not found at this Tm | N.d. | 0.02 |
mm4_intergenic_Ntrk2|Agtpbp1_chr13_59360879_R | Primer3: not found at this Tm | N.d. | 0.02 |
mm4_intergenic_Zpld1|Mir5118_chr16_55357462_F | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intergenic_Zpld1|Mir5118_chr16_55357462_R | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intron_Gm20442_chr17_35743837_F | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intron_Gm20442_chr17_35743837_R | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intergenic_Polb|Ikbkb_chr8_22656565_F | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intergenic_Polb|Ikbkb_chr8_22656565_R | Primer3: not found at this Tm | N.d. | 0.01 |
mm4_intergenic_Gm12741|3110021N24Rik/Zfyve9_chr4_108733014_F | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Gm12741|3110021N24Rik/Zfyve9_chr4_108733014_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_Med16_chr10_79907907 |
CACTTCAGTCTGGGCCTCAGTCTACACTCTAGTTCAGTCTCCAAGCCTTGCCTGGGACCTAGTAGCTTAGACCCCGCAGC AGCCCATGCCACACAATACT |
mm4_exon_Palld_chr8_61720535 |
ACCATTCACACTTTGCCCCTCCACAGCCTCCTTACCTTCAATGAACACCTCTGCAGATGTGCTGTCTGAGCCACTGGGGT TTGTAGCCAGACACGTGTAGCGACCTGTGTCGTCCTCAAAGGCCTCAGCAATGACCA |
mm4_intergenic_Actb|Fscn1_chr5_142936443 |
GGACAATTACAGGCTGGGCTCAGCTCTGGGGTTTACTTTTCTTAGGGACGGAAGAAAAGAGGGCCCCACTCTTTGAATTC CACGTAGTTCAGACCTCGAATCGCCTCTCCGGGATAGGCTGCTATGT |
mm4_intron_Nrg2_chr18_36118511 |
CCTCACAGAGACCTCCAGGACACAAACTCCCTGAAGCTCAGACACATAATCACAAGCACATGCACTCAAATCACACATTT CCAAAAAGGGCCGTAAACATGCCCTTTTCCCTAGGC |
mm4_intron_Cdc14a_chr3_116307187 |
ACTAGCACAGTAGCTTCCAGCTCCCCAAATGCTCAGACTCCGAATCCATCGTAAGTCATTTCCTGCTACTTCAGAGAAAG CAGAGAACAATAGGAAAAAAAAAAAAAAAAAGCATGAGTTTGGGATCAGACA |
mm4_intergenic_Dancr/Snora26|Rasl11b_chr5_74097935 |
GCTCGCTTTTCACACCAAGGGTAGGCAGCGGTGCTTTAGGTCCGGGTGTTGGGGTTAGTTTATTAACCCGAGTGACTCAG ACAACGAAACAACCTCGGGACCCACAGAAGCATCAAGGGTTCTCGGACA |
mm4_intergenic_Gm5205|Sh2d4b_chr14_40693943 |
GGTAGTCAGGCTTCAGGCATACATATATTAACATCGATTTCAGATATATTCACTCTACATTGGAACTAAAAACACCCAGT AGCCCAGGGACAGAATCACAGAAAAGGCAAAGTCACCTGAAGACA |
mm4_intron_Pitpnc1_chr11_107249066 |
CTGTTTCGCTGGCACTGATGGACCTGGGCATGTGACTTGGCACCAAGAGCTTTCTGAATTCCATAGGCAGCCAGGACTCT GTGTCTGTGCTGCTTGGTAGAGGTGCTCTCTCTGACCTCCCAATTGTGTGTGTGTGCA |
mm4_intergenic_Mrps28|Gm15467_chr3_8850429 |
GGATAGCCTGCAGAGACGACACAGCAACAGAGAAAGAGGTGTCTGAGCTACTGAGGCAATCTCCTCCTAGTGACTGCTGA AGGCTTCATCAGAGGGTGGGTC |
mm4_intergenic_Hipk3|Gm13870_chr2_104508381 |
TCACTGCTTGCCACTTCTGAGTGGCTCAGTAGNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNAAAAGGAGAAGGGTCGGCTG |
mm4_intron_Nos1ap_chr1_170423646 |
CCCAGAAGGACTTAGCAGAGCATTAAGGTAATCTCACTGTTCACTGTCCATATGAGAAGAACAAACTATCCCAGTCGCCC AGACGCAGAATCTGGCAATTCTGTTCCTTTTCTTCTTTACCTCCTTTCCAGCCTCCA |
mm4_intron_Slc9a9_chr9_95193065 |
GGAAGGCATGTCCACACAGAGCCATCAAGCCATGCACTTCCTGGTAGCTCAGCCAGTGAATCTCCAGGAGGTGATGGCCA CAGTGTGGATTTTTCCCATCTCTCTTTTCCCCAAGGCCATCTCTAGG |
mm4_intergenic_Gm23653|Nnmt_chr9_48580114 |
TGTCAGAGATTGTCCATGTTGTGATTCAGTGTCTGATCTTATAGGTTCAGAAACTTGATTTATCATTACCTTCAAGGTGG ATATAGTCAGTGATTCACTGACAAAACAAGTAAGAGACTAAAAGCAGACTCAGCT |
mm4_intergenic_Gm5973|Gm9915_chr1_42186377 |
ATGGACTCACCTGGACCTGACCAGCCCCTGTCAACAGTAAGAGCCAGCAAATAAAGCTTATTGTAGTGACTGCTAATATG ATTCCTTTTCTGATCTACTAGGGCAGATAGGAATAACCATTTCAACCCTGGAGCA |
mm4_intergenic_Fto|Irx3_chr8_91711866 |
CTCCTGATTCCTGCTGCGATGGGCTGTTCCTGGGCTGTTCTTGTCTCTGAGATTCCCAGGTAGCTCAGACCCCAGATCAG TGGGAACGGAATCTCTGCCCGAT |
mm4_intergenic_Gm24069|Gm24434_chr12_39521785 |
GCTGGTGCTGAGAATATAGTGGTCTCAGCATGGGAGTCTGTGGCTGATCTACTGGGACAATGTTTGATATTGTGAGAAAG TTGTATTTTGAGTCA |