← return to the list of all guides
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_52313974_F | Primer3: not found at this Tm | N.d. | 1.00 |
ontarget_mm0_exon_5730457N03Rik/Evx1_chr6_52313974_R | Primer3: not found at this Tm | N.d. | 1.00 |
mm4_intergenic_Gm13462|Lrp1b_chr2_40982455_F | Primer3: not found at this Tm | N.d. | 0.34 |
mm4_intergenic_Gm13462|Lrp1b_chr2_40982455_R | Primer3: not found at this Tm | N.d. | 0.34 |
mm4_exon_St3gal2_chr8_110962292_F | Primer3: not found at this Tm | N.d. | 0.33 |
mm4_exon_St3gal2_chr8_110962292_R | Primer3: not found at this Tm | N.d. | 0.33 |
mm3_intergenic_4930428D18Rik|Gm5793_chrX_76481467_F | Primer3: not found at this Tm | N.d. | 0.20 |
mm3_intergenic_4930428D18Rik|Gm5793_chrX_76481467_R | Primer3: not found at this Tm | N.d. | 0.20 |
mm4_intergenic_Gm22673|Gm23059_chr12_60309499_F | Primer3: not found at this Tm | N.d. | 0.19 |
mm4_intergenic_Gm22673|Gm23059_chr12_60309499_R | Primer3: not found at this Tm | N.d. | 0.19 |
mm4_intron_Ttl_chr2_129071597_F | Primer3: not found at this Tm | N.d. | 0.18 |
mm4_intron_Ttl_chr2_129071597_R | Primer3: not found at this Tm | N.d. | 0.18 |
mm4_intergenic_Zscan20|Tlr12_chr4_128610887_F | Primer3: not found at this Tm | N.d. | 0.17 |
mm4_intergenic_Zscan20|Tlr12_chr4_128610887_R | Primer3: not found at this Tm | N.d. | 0.17 |
mm4_intron_G930045G22Rik_chr6_50849884_F | Primer3: not found at this Tm | N.d. | 0.15 |
mm4_intron_G930045G22Rik_chr6_50849884_R | Primer3: not found at this Tm | N.d. | 0.15 |
mm4_intergenic_Smad2|Skor2_chr18_76654132_F | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intergenic_Smad2|Skor2_chr18_76654132_R | Primer3: not found at this Tm | N.d. | 0.13 |
mm4_intergenic_Srfbp1|Gm27026_chr18_52465935_F | Primer3: not found at this Tm | N.d. | 0.12 |
mm4_intergenic_Srfbp1|Gm27026_chr18_52465935_R | Primer3: not found at this Tm | N.d. | 0.12 |
mm4_intergenic_Hmgb1-ps9|Pik3r1_chr13_101441368_F | TCGTCGGCAGCGTCCCACAGTATCTCCACAACAGCT | 60.0 | 0.11 |
mm4_intergenic_Hmgb1-ps9|Pik3r1_chr13_101441368_R | GTCTCGTGGGCTCGGCCCGACTCTCTCCCACTCAT | 60.3 | 0.11 |
mm4_intergenic_1700123L14Rik|Gm22971_chr6_96330769_F | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_intergenic_1700123L14Rik|Gm22971_chr6_96330769_R | Primer3: not found at this Tm | N.d. | 0.09 |
mm4_intergenic_Gm11467|Gm11468_chr2_166232722_F | Primer3: not found at this Tm | N.d. | 0.08 |
mm4_intergenic_Gm11467|Gm11468_chr2_166232722_R | Primer3: not found at this Tm | N.d. | 0.08 |
mm4_intergenic_Fam83b|RP23-264P8.1_chr9_76612324_F | Primer3: not found at this Tm | N.d. | 0.07 |
mm4_intergenic_Fam83b|RP23-264P8.1_chr9_76612324_R | Primer3: not found at this Tm | N.d. | 0.07 |
mm4_intergenic_Gm22444|mmu-mir-6238_chr7_53702076_F | Primer3: not found at this Tm | N.d. | 0.07 |
mm4_intergenic_Gm22444|mmu-mir-6238_chr7_53702076_R | Primer3: not found at this Tm | N.d. | 0.07 |
mm4_intergenic_Fbxo2|Ptchd2_chr4_148185204_F | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_intergenic_Fbxo2|Ptchd2_chr4_148185204_R | Primer3: not found at this Tm | N.d. | 0.05 |
mm4_exon_Tet3_chr6_83370517_F | TCGTCGGCAGCGTCTTCTCTGGCGTGCTCAGTTT | 59.8 | 0.01 |
mm4_exon_Tet3_chr6_83370517_R | GTCTCGTGGGCTCGGCACAGCATTCCCCAGAGAGG | 60.1 | 0.01 |
mm4_intron_Prkg2_chr5_99036892_F | TCGTCGGCAGCGTCGTGTGGGAGAAGTGCCCAC | 60.6 | 0.00 |
mm4_intron_Prkg2_chr5_99036892_R | GTCTCGTGGGCTCGGCTGCGCACCTGGAAAGTTAC | 59.4 | 0.00 |
mm4_intron_Abcc4_chr14_118665668_F | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intron_Abcc4_chr14_118665668_R | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Prss30|Prss22_chr17_23991201_F | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Prss30|Prss22_chr17_23991201_R | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Il34|Gm15894_chr8_110750509_F | TCGTCGGCAGCGTCTGGTGATGTACGTTACGGGG | 59.4 | 0.00 |
mm4_intergenic_Il34|Gm15894_chr8_110750509_R | GTCTCGTGGGCTCGGTTTTCAAAGCACGGGAGGTT | 58.2 | 0.00 |
mm4_intergenic_Gm26312|Fhl1_chrX_56731331_F | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Gm26312|Fhl1_chrX_56731331_R | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Amn1|2810474O19Rik_chr6_149294974_F | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_intergenic_Amn1|2810474O19Rik_chr6_149294974_R | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_exon_Sowahb/9330159N05Rik_chr5_93043987_F | TCGTCGGCAGCGTCCTGGAGGAAGTTGTCTGGCA | 59.6 | 0.00 |
mm4_exon_Sowahb/9330159N05Rik_chr5_93043987_R | GTCTCGTGGGCTCGGAACCATTCAACCCAGCAGCT | 60.1 | 0.00 |
mm4_exon_Cspg5_chr9_110245092_F | Primer3: not found at this Tm | N.d. | 0.00 |
mm4_exon_Cspg5_chr9_110245092_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_intergenic_Hmgb1-ps9|Pik3r1_chr13_101441368 |
CCACAGTATCTCCACAACAGCTATGGAAACATGAAAACCTTATCTACTCAGGGGCTCAGGCAATTGTTGGATAGAATTAT AGCCTGTTGTGAATGGTCTCCCAATCCCAGATGAGTGGGAGAGAGTCGGG |
mm4_exon_Tet3_chr6_83370517 |
TTCTCTGGCGTGCTCAGTTTCTCCTTCTGCAGCTTCTTCTTCTCGGCCGCCGCCTTCCTGGCTTCCAGCTGCCGTTGGCG GCAGGACTTGGCAGGCTCAGGCAGCCGCCGGACCTCTCTGGGGAATGCTGTG |
mm4_intron_Prkg2_chr5_99036892 |
GTGTGGGAGAAGTGCCCACCGCGCAACGCACGCCCAGTGGCACCCGCGTGTCCCCACTCCCGTCCCTGGTCGCGAGCCCC ACGGGTGCCCGAGCCCCCCAGCCTCACCAGGCCGCCGGGTAACTTTCCAGGTGCGCAG |
mm4_intergenic_Il34|Gm15894_chr8_110750509 |
TGGTGATGTACGTTACGGGGGGAATGTTTAAGATATGTTCAACGGCCTTCCTAAGGAAGCCTGAGCCAGCGATATCAGCA GCTTAAAGGATGACTTCAAGTTTATTAATTAACCTCCCGTGCTTTGAAAA |
mm4_exon_Sowahb/9330159N05Rik_chr5_93043987 |
CTGGAGGAAGTTGTCTGGCAACACCGACCAGGCGCGAGTGGGCGCCTGGATCTCGGGCTCAGGCACCTGCAGGTATCTGG CTACCCACTCTTGAGTGCGCTGGTGCTGTTGCTGCAGCTGCTGGGTTGAATGGTT |