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Contents:

PCR primers for off-targets of CAACGGCTCCGGCATGTGCA AGG

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.

NamePrimer Sequence Tm CFD Score
ontarget_mm0__chr7 5.53 Mbp_chr7_5529602_F TCGTCGGCAGCGTCCACGATGGAGGGGAAGACG 59.8 1.00
ontarget_mm0__chr7 5.53 Mbp_chr7_5529602_R GTCTCGTGGGCTCGGGCTATTCTCGCAGCTCACCA 60.1 1.00
mm0__chr5 77.78 Mbp_chr5_77784918_F TCGTCGGCAGCGTCATATCACCGCGCTCGTCATT 59.9 1.00
mm0__chr5 77.78 Mbp_chr5_77784918_R GTCTCGTGGGCTCGGAACGATGGAGGGGAAGACTG 59.0 1.00
mm1__chr22 15.72 Mbp_chr22_15721518_F TCGTCGGCAGCGTCATACTGCCGTGCTCGTCATT 59.8 0.94
mm1__chr22 15.72 Mbp_chr22_15721518_R GTCTCGTGGGCTCGGATCATGTTCTGGTGCCTGGG 60.0 0.94
mm1__chr2 131.63 Mbp_chr2_131626170_F TCGTCGGCAGCGTCATACCGCTGTGCTCGTCATT 59.8 0.94
mm1__chr2 131.63 Mbp_chr2_131626170_R GTCTCGTGGGCTCGGCTTCTGATGCATGCCCTCCA 60.1 0.94
mm1__chr2 131.26 Mbp_chr2_131263587_F TCGTCGGCAGCGTCTGGATGATGATACCGCCGTG 59.9 0.94
mm1__chr2 131.26 Mbp_chr2_131263587_R GTCTCGTGGGCTCGGGATGGAAGGGAAGACAGCCC 60.1 0.94
mm1__chr2 130.66 Mbp_chr2_130656781_F TCGTCGGCAGCGTCATACCGCTGTGCTCGTCATT 59.8 0.94
mm1__chr2 130.66 Mbp_chr2_130656781_R GTCTCGTGGGCTCGGGATGGAAGGGAAGACAGCCC 60.1 0.94
mm1__chr2 130.46 Mbp_chr2_130463888_F TCGTCGGCAGCGTCGATGGAAGGGAAGACAGCCC 60.1 0.94
mm1__chr2 130.46 Mbp_chr2_130463888_R GTCTCGTGGGCTCGGATGATGATACCGCCGTGCTC 60.3 0.94
mm1__chr2 130.08 Mbp_chr2_130075316_F TCGTCGGCAGCGTCGATGGAAGGGAAGACAGCCC 60.1 0.94
mm1__chr2 130.08 Mbp_chr2_130075316_R GTCTCGTGGGCTCGGATACCGCTGTGCTCGTCATT 59.8 0.94
mm1__chr14 19.40 Mbp_chr14_19402853_F TCGTCGGCAGCGTCATCATGTTCTGGTGCCTGGG 60.0 0.94
mm1__chr14 19.40 Mbp_chr14_19402853_R GTCTCGTGGGCTCGGATACTGCCGTGCTCGTCATT 59.8 0.94
mm1__chr14 19.00 Mbp_chr14_18998899_F TCGTCGGCAGCGTCATACTGCCGTGCTCGTCATT 59.8 0.94
mm1__chr14 19.00 Mbp_chr14_18998899_R GTCTCGTGGGCTCGGATCATGTTCTGGTGCCTGGG 60.0 0.94
mm1__chr9 100.73 Mbp_chr9_100731271_F TCGTCGGCAGCGTCGCCAGCTCACCATGGATGAT 60.1 0.86
mm1__chr9 100.73 Mbp_chr9_100731271_R GTCTCGTGGGCTCGGCTAGGGAGCTCCACGATGGA 60.4 0.86
mm1__chr1 223.86 Mbp_chr1_223864796_F TCGTCGGCAGCGTCCATGGAAGGGAAGATGGCCC 60.4 0.86
mm1__chr1 223.86 Mbp_chr1_223864796_R GTCTCGTGGGCTCGGGCCAGCTCACCATGGATGAT 60.1 0.86
mm2__chr22 41.08 Mbp_chr22_41075184_F TCGTCGGCAGCGTCATGCCCACCATCATGACCTG 60.1 0.81
mm2__chr22 41.08 Mbp_chr22_41075184_R GTCTCGTGGGCTCGGTATTGCTGCGCTCGTTGTTG 59.8 0.81
mm1__chr19 9.52 Mbp_chr19_9518181_F TCGTCGGCAGCGTCGCTGTCAGCTCACCATGGAT 60.1 0.80
mm1__chr19 9.52 Mbp_chr19_9518181_R GTCTCGTGGGCTCGGCCACGATGGAGGGGAAGATG 59.8 0.80
mm2__chrX 46.29 Mbp_chrX_46289073_F TCGTCGGCAGCGTCTGGAGGGGAAGACAACTGGG 61.1 0.79
mm2__chrX 46.29 Mbp_chrX_46289073_R GTCTCGTGGGCTCGGCTCCATCACCAGTCCACACC 60.0 0.79
mm3__chr5 80.30 Mbp_chr5_80302792_F TCGTCGGCAGCGTCATGATGAAGGGGAAGGCAGC 60.1 0.75
mm3__chr5 80.30 Mbp_chr5_80302792_R GTCTCGTGGGCTCGGAGCTCACCATGCATGACGAT 59.8 0.75
mm2__chr18 62.44 Mbp_chr18_62442040_F Primer3: not found at this Tm N.d. 0.75
mm2__chr18 62.44 Mbp_chr18_62442040_R Primer3: not found at this Tm N.d. 0.75
mm2__chr15 43.99 Mbp_chr15_43990129_F TCGTCGGCAGCGTCCATCGTCACCTGCAAAGCTG 59.8 0.75
mm2__chr15 43.99 Mbp_chr15_43990129_R GTCTCGTGGGCTCGGATCACCATGCTTGTCGTCGA 59.7 0.75
mm4__chr13 47.12 Mbp_chr13_47118582_F Primer3: not found at this Tm N.d. 0.64
mm4__chr13 47.12 Mbp_chr13_47118582_R Primer3: not found at this Tm N.d. 0.64
mm3__chr11 1.80 Mbp_chr11_1804040_F TCGTCGGCAGCGTCCACGATGGAGGGGAAGACG 59.8 0.55
mm3__chr11 1.80 Mbp_chr11_1804040_R GTCTCGTGGGCTCGGCCCAGCTCACCATGGATGAT 59.5 0.55
mm2__chr5 98.34 Mbp_chr5_98338775_F TCGTCGGCAGCGTCGCAGCTAGCTCACCATGGAT 59.8 0.47
mm2__chr5 98.34 Mbp_chr5_98338775_R GTCTCGTGGGCTCGGGCATTGTCACCTGTGAAGCC 59.7 0.47
mm4__chr8 35.75 Mbp_chr8_35745491_F TCGTCGGCAGCGTCCCAAGGAGCCTTAAATGCCC 58.8 0.41
mm4__chr8 35.75 Mbp_chr8_35745491_R GTCTCGTGGGCTCGGTGTCATTGCATCAGTGTACGC 59.2 0.41
mm4__chr8 107.80 Mbp_chr8_107798187_F Primer3: not found at this Tm N.d. 0.35
mm4__chr8 107.80 Mbp_chr8_107798187_R Primer3: not found at this Tm N.d. 0.35
mm4__chr1 2.17 Mbp_chr1_2172168_F TCGTCGGCAGCGTCGGACGGGCACATCAAGCT 60.0 0.30
mm4__chr1 2.17 Mbp_chr1_2172168_R GTCTCGTGGGCTCGGCTTGTAGACACTACCCCGCG 60.1 0.30
mm4__chr12 27.78 Mbp_chr12_27780428_F TCGTCGGCAGCGTCGAGCAGAGCGACTACTTCCG 60.2 0.30
mm4__chr12 27.78 Mbp_chr12_27780428_R GTCTCGTGGGCTCGGGCGTTGATGAAGTCCAGCAC 59.8 0.30
mm4__chr20 21.04 Mbp_chr20_21044449_F Primer3: not found at this Tm N.d. 0.27
mm4__chr20 21.04 Mbp_chr20_21044449_R Primer3: not found at this Tm N.d. 0.27
mm3__chr6 88.28 Mbp_chr6_88276931_F TCGTCGGCAGCGTCCCCACAATGGAGGGGAAGAC 60.0 0.21
mm3__chr6 88.28 Mbp_chr6_88276931_R GTCTCGTGGGCTCGGGCCAGCTCACCATGGATGAT 60.1 0.21
mm2__chr5 61.83 Mbp_chr5_61832990_F TCGTCGGCAGCGTCGACGTTGTGTCCAGCAAAGC 60.3 0.20
mm2__chr5 61.83 Mbp_chr5_61832990_R GTCTCGTGGGCTCGGTCCTTGGCCAAAGCAGTGAA 60.1 0.20
mm2__chr20 1.16 Mbp_chr20_1160615_F TCGTCGGCAGCGTCGCTCCTCCACCGATCACAAT 59.8 0.20
mm2__chr20 1.16 Mbp_chr20_1160615_R GTCTCGTGGGCTCGGGGGAAACATGGTTCCAGGGG 60.6 0.20
mm2__chr1 237.93 Mbp_chr1_237926862_F TCGTCGGCAGCGTCCTCCTCCACCAGTCACGATG 59.8 0.20
mm2__chr1 237.93 Mbp_chr1_237926862_R GTCTCGTGGGCTCGGTGGAGGGAAACACAGCTTGG 60.1 0.20
mm2__chr1 103.57 Mbp_chr1_103569584_F Primer3: not found at this Tm N.d. 0.20
mm2__chr1 103.57 Mbp_chr1_103569584_R Primer3: not found at this Tm N.d. 0.20
mm1__chr17 81.51 Mbp_chr17_81512299_F TCGTCGGCAGCGTCCCATCCACTCACCTGGTGTC 60.0 0.20
mm1__chr17 81.51 Mbp_chr17_81512299_R GTCTCGTGGGCTCGGATGGAAGAAGAGATCGCCGC 60.2 0.20
mm3__chr8 84.95 Mbp_chr8_84948621_F TCGTCGGCAGCGTCCTGCTTTGCCACTGGTTCAC 59.9 0.19
mm3__chr8 84.95 Mbp_chr8_84948621_R GTCTCGTGGGCTCGGGAAAGGAAGAAGGCCTGGGG 60.3 0.19
mm3__chr5 15.45 Mbp_chr5_15450731_F Primer3: not found at this Tm N.d. 0.19
mm3__chr5 15.45 Mbp_chr5_15450731_R Primer3: not found at this Tm N.d. 0.19
mm2__chrX 53.14 Mbp_chrX_53143858_F TCGTCGGCAGCGTCGATGGAGGGGAACACAGCTC 60.1 0.19
mm2__chrX 53.14 Mbp_chrX_53143858_R GTCTCGTGGGCTCGGTGCTCTTCCACAGGTTGCAA 60.1 0.19
mm4__chr5 138.28 Mbp_chr5_138283423_F TCGTCGGCAGCGTCTAGGGCAAGAGCATGTGCAA 59.9 0.18
mm4__chr5 138.28 Mbp_chr5_138283423_R GTCTCGTGGGCTCGGAAAGTGAGTGGGAGGGAAGG 58.9 0.18
mm3__chr12 120.35 Mbp_chr12_120353789_F Primer3: not found at this Tm N.d. 0.17
mm3__chr12 120.35 Mbp_chr12_120353789_R Primer3: not found at this Tm N.d. 0.17
mm4__chr12 110.80 Mbp_chr12_110803125_F Primer3: not found at this Tm N.d. 0.17
mm4__chr12 110.80 Mbp_chr12_110803125_R Primer3: not found at this Tm N.d. 0.17
mm4__chr3 180.82 Mbp_chr3_180820575_F TCGTCGGCAGCGTCAATGATATCGCCACGCTCGT 59.9 0.17
mm4__chr3 180.82 Mbp_chr3_180820575_R GTCTCGTGGGCTCGGTTCTTCTGACCCATGCCCAC 59.9 0.17
mm4__chr3 49.17 Mbp_chr3_49171474_F TCGTCGGCAGCGTCCCAGTGTCCAAGTGTCCGTT 60.1 0.16
mm4__chr3 49.17 Mbp_chr3_49171474_R GTCTCGTGGGCTCGGGGAAACCTCCTCTCGCCTTA 58.8 0.16
mm3__chr2 73.90 Mbp_chr2_73901346_F TCGTCGGCAGCGTCGTGTGAAGAGGAGACCACCG 60.0 0.16
mm3__chr2 73.90 Mbp_chr2_73901346_R GTCTCGTGGGCTCGGCCCACAATGGAGGGGAAGAC 60.0 0.16
mm4__chr3 59.58 Mbp_chr3_59579128_F TCGTCGGCAGCGTCTGCACTGATGTGAAGCCCAT 59.9 0.14
mm4__chr3 59.58 Mbp_chr3_59579128_R GTCTCGTGGGCTCGGAGCATTCTGATCCGTTTGGC 58.9 0.14
mm4__chr12 14.37 Mbp_chr12_14365639_F Primer3: not found at this Tm N.d. 0.12
mm4__chr12 14.37 Mbp_chr12_14365639_R Primer3: not found at this Tm N.d. 0.12
mm4__chr12 132.07 Mbp_chr12_132067382_F TCGTCGGCAGCGTCCAGGACAGACCGTGGTGG 59.6 0.12
mm4__chr12 132.07 Mbp_chr12_132067382_R GTCTCGTGGGCTCGGCCGGTACCTTCTGCTGGAC 59.7 0.12
mm4__chr2 67.82 Mbp_chr2_67823888_F TCGTCGGCAGCGTCTTTGGCAGCCTCATGGTTCA 60.1 0.12
mm4__chr2 67.82 Mbp_chr2_67823888_R GTCTCGTGGGCTCGGAAGAGCACTGTGGGTGATGG 59.9 0.12
mm1__chr9 6.83 Mbp_chr9_6834487_F TCGTCGGCAGCGTCCCGCTACCAGCTCACCATAG 59.9 0.12
mm1__chr9 6.83 Mbp_chr9_6834487_R GTCTCGTGGGCTCGGACACGATGAAGGGGAAGACG 59.7 0.12
mm4__chr2 57.76 Mbp_chr2_57764984_F TCGTCGGCAGCGTCAGATTGCTGGCTCGTCAATGA 60.0 0.11
mm4__chr2 57.76 Mbp_chr2_57764984_R GTCTCGTGGGCTCGGCTTCTTGCCCATGCCCACTA 60.0 0.11
mm4__chr3 89.59 Mbp_chr3_89589793_F Primer3: not found at this Tm N.d. 0.11
mm4__chr3 89.59 Mbp_chr3_89589793_R Primer3: not found at this Tm N.d. 0.11
mm4__chr2 211.78 Mbp_chr2_211778667_F TCGTCGGCAGCGTCGGAACCGGCTGGACAGTTTA 59.9 0.11
mm4__chr2 211.78 Mbp_chr2_211778667_R GTCTCGTGGGCTCGGGGAGCGAGAAGATGGTCAGG 59.8 0.11
mm4__chr2 202.62 Mbp_chr2_202619468_F TCGTCGGCAGCGTCGGAGCGAGAAGATGGTCAGG 59.8 0.11
mm4__chr2 202.62 Mbp_chr2_202619468_R GTCTCGTGGGCTCGGTCAATAGTAGAAGCCGGCGC 60.2 0.11
mm4__chr1 106.81 Mbp_chr1_106805614_F TCGTCGGCAGCGTCCACCTGCTAAGTGGAGCGAG 60.4 0.11
mm4__chr1 106.81 Mbp_chr1_106805614_R GTCTCGTGGGCTCGGGCTGGCGCTTGAATCAGCTA 61.0 0.11
mm4__chr13 111.35 Mbp_chr13_111346406_F TCGTCGGCAGCGTCATCAGGGACAGCCAGCCAT 61.0 0.11
mm4__chr13 111.35 Mbp_chr13_111346406_R GTCTCGTGGGCTCGGTTCCTTGGCAGCTCTGTCTG 59.9 0.11
mm3__chr9 135.73 Mbp_chr9_135728729_F TCGTCGGCAGCGTCGCCCAGCTTGGAGAGACATC 60.4 0.09
mm3__chr9 135.73 Mbp_chr9_135728729_R GTCTCGTGGGCTCGGTAACCAGACCCGGCAAAACA 59.8 0.09
mm3__chr5 131.66 Mbp_chr5_131660060_F TCGTCGGCAGCGTCATGCCCATCATCACACCCTG 60.1 0.09
mm3__chr5 131.66 Mbp_chr5_131660060_R GTCTCGTGGGCTCGGATTGCTGTGCTTGTCGTCGA 60.6 0.09
mm3__chr22 35.54 Mbp_chr22_35535674_F Primer3: not found at this Tm N.d. 0.09
mm3__chr22 35.54 Mbp_chr22_35535674_R Primer3: not found at this Tm N.d. 0.09
mm2__chr6 46.21 Mbp_chr6_46205009_F TCGTCGGCAGCGTCATGGAAGAAGAGATCCCCGC 59.2 0.08
mm2__chr6 46.21 Mbp_chr6_46205009_R GTCTCGTGGGCTCGGATATGGCTCGGAGGGCATTG 59.9 0.08
mm4__chr5 135.07 Mbp_chr5_135066866_F TCGTCGGCAGCGTCAGGGAGGCTGATGTGAAAGC 60.0 0.08
mm4__chr5 135.07 Mbp_chr5_135066866_R GTCTCGTGGGCTCGGTCCTGGGGTTGGTGTATGGA 60.1 0.08
mm4__chr17 16.47 Mbp_chr17_16466367_F TCGTCGGCAGCGTCCTTTCAGACTCGGCACCCAT 60.0 0.08
mm4__chr17 16.47 Mbp_chr17_16466367_R GTCTCGTGGGCTCGGCAGGGACCACACCTGGATCT 60.9 0.08
mm4__chrX 52.96 Mbp_chrX_52959493_F Primer3: not found at this Tm N.d. 0.07
mm4__chrX 52.96 Mbp_chrX_52959493_R Primer3: not found at this Tm N.d. 0.07
mm4__chrX 52.90 Mbp_chrX_52896297_F Primer3: not found at this Tm N.d. 0.07
mm4__chrX 52.90 Mbp_chrX_52896297_R Primer3: not found at this Tm N.d. 0.07
mm3__chrY 18.15 Mbp_chrY_18147915_F Primer3: not found at this Tm N.d. 0.07
mm3__chrY 18.15 Mbp_chrY_18147915_R Primer3: not found at this Tm N.d. 0.07
mm3__chrY 17.76 Mbp_chrY_17758070_F TCGTCGGCAGCGTCAACGGTGGAGGGAAACATGG 60.2 0.07
mm3__chrY 17.76 Mbp_chrY_17758070_R GTCTCGTGGGCTCGGTCCTCTGTCCCACTCCTCTG 59.9 0.07
mm4__chr4 180.31 Mbp_chr4_180311834_F Primer3: not found at this Tm N.d. 0.07
mm4__chr4 180.31 Mbp_chr4_180311834_R Primer3: not found at this Tm N.d. 0.07
mm4__chr6 169.65 Mbp_chr6_169648288_F TCGTCGGCAGCGTCTCTACAGATGCCTTCCGTGC 59.8 0.07
mm4__chr6 169.65 Mbp_chr6_169648288_R GTCTCGTGGGCTCGGCTGGCTCTTCTGCATTTGGC 59.8 0.07
mm4__chr3 46.88 Mbp_chr3_46875620_F TCGTCGGCAGCGTCCACCACCTCCCTGGATTTCC 60.0 0.06
mm4__chr3 46.88 Mbp_chr3_46875620_R GTCTCGTGGGCTCGGTGCAGCGTCGTAACAGTTCT 59.9 0.06
mm4__chr10 5.59 Mbp_chr10_5585443_F TCGTCGGCAGCGTCTTCTGAGGACTCACAGCTGC 59.6 0.05
mm4__chr10 5.59 Mbp_chr10_5585443_R GTCTCGTGGGCTCGGGAAGGGAGACAAGGGTGGTG 59.9 0.05
mm3__chrX 75.76 Mbp_chrX_75757960_F Primer3: not found at this Tm N.d. 0.05
mm3__chrX 75.76 Mbp_chrX_75757960_R Primer3: not found at this Tm N.d. 0.05
mm4__chr3 169.77 Mbp_chr3_169769465_F Primer3: not found at this Tm N.d. 0.05
mm4__chr3 169.77 Mbp_chr3_169769465_R Primer3: not found at this Tm N.d. 0.05
mm4__chr19 31.57 Mbp_chr19_31566196_F TCGTCGGCAGCGTCTTCACCAGAGCCCAACGTAC 59.9 0.05
mm4__chr19 31.57 Mbp_chr19_31566196_R GTCTCGTGGGCTCGGCCCTACCCTATGCCCCAGTA 59.8 0.05
mm4__chr8 138.19 Mbp_chr8_138192551_F Primer3: not found at this Tm N.d. 0.05
mm4__chr8 138.19 Mbp_chr8_138192551_R Primer3: not found at this Tm N.d. 0.05
mm4__chr20 59.74 Mbp_chr20_59737337_F TCGTCGGCAGCGTCTCTGTGGGGATCCTGGTTCA 60.1 0.04
mm4__chr20 59.74 Mbp_chr20_59737337_R GTCTCGTGGGCTCGGTGCAGCTGTATCTCCAGTGC 60.1 0.04
mm4__chr1 248.18 Mbp_chr1_248176264_F TCGTCGGCAGCGTCCAGTGACTTCTCAATTTCCGACC 59.5 0.04
mm4__chr1 248.18 Mbp_chr1_248176264_R GTCTCGTGGGCTCGGTCAGATTTCATGACAGTGTGGTG 58.9 0.04
mm3__chrX 122.37 Mbp_chrX_122365273_F Primer3: not found at this Tm N.d. 0.04
mm3__chrX 122.37 Mbp_chrX_122365273_R Primer3: not found at this Tm N.d. 0.04
mm4__chrX 152.91 Mbp_chrX_152906792_F TCGTCGGCAGCGTCGTCTGCTCAGCTCCTGGATG 60.1 0.02
mm4__chrX 152.91 Mbp_chrX_152906792_R GTCTCGTGGGCTCGGGGCTTGTCTCAGTGCATGTG 59.4 0.02
mm4__chr1 226.77 Mbp_chr1_226771924_F Primer3: not found at this Tm N.d. 0.02
mm4__chr1 226.77 Mbp_chr1_226771924_R Primer3: not found at this Tm N.d. 0.02
mm4__chr3 74.00 Mbp_chr3_74002249_F TCGTCGGCAGCGTCAGAGCAGCCATGAACTGACA 59.3 0.02
mm4__chr3 74.00 Mbp_chr3_74002249_R GTCTCGTGGGCTCGGTTGAGCAAAGTGTTGGTGCC 59.5 0.02
mm4__chr18 10.33 Mbp_chr18_10330721_F Primer3: not found at this Tm N.d. 0.01
mm4__chr18 10.33 Mbp_chr18_10330721_R Primer3: not found at this Tm N.d. 0.01
mm4__chr2 205.39 Mbp_chr2_205387855_F TCGTCGGCAGCGTCTGGGTAATGGTTGCCAGTCC 59.9 0.01
mm4__chr2 205.39 Mbp_chr2_205387855_R GTCTCGTGGGCTCGGAACTGGCAGCCTGTGTTTCT 60.1 0.01
mm4__chr9 69.43 Mbp_chr9_69425355_F Primer3: not found at this Tm N.d. 0.01
mm4__chr9 69.43 Mbp_chr9_69425355_R Primer3: not found at this Tm N.d. 0.01
mm4__chr16 28.32 Mbp_chr16_28320766_F Primer3: not found at this Tm N.d. 0.01
mm4__chr16 28.32 Mbp_chr16_28320766_R Primer3: not found at this Tm N.d. 0.01
mm4__chr19 45.03 Mbp_chr19_45033006_F Primer3: not found at this Tm N.d. 0.01
mm4__chr19 45.03 Mbp_chr19_45033006_R Primer3: not found at this Tm N.d. 0.01
mm4__chr12 1.57 Mbp_chr12_1570341_F Primer3: not found at this Tm N.d. 0.00
mm4__chr12 1.57 Mbp_chr12_1570341_R Primer3: not found at this Tm N.d. 0.00
mm4__chr20 56.27 Mbp_chr20_56271924_F Primer3: not found at this Tm N.d. 0.00
mm4__chr20 56.27 Mbp_chr20_56271924_R Primer3: not found at this Tm N.d. 0.00
mm4__chr19 42.04 Mbp_chr19_42042571_F Primer3: not found at this Tm N.d. 0.00
mm4__chr19 42.04 Mbp_chr19_42042571_R Primer3: not found at this Tm N.d. 0.00
mm4__chr7 157.09 Mbp_chr7_157087829_F TCGTCGGCAGCGTCTGGCATACAGTAGGGCTTGC 60.1 0.00
mm4__chr7 157.09 Mbp_chr7_157087829_R GTCTCGTGGGCTCGGCTGCCTTCCGGATATCCACG 60.3 0.00
mm4__chr7 101.09 Mbp_chr7_101088299_F Primer3: not found at this Tm N.d. 0.00
mm4__chr7 101.09 Mbp_chr7_101088299_R Primer3: not found at this Tm N.d. 0.00
mm4__chr4 53.48 Mbp_chr4_53477346_F Primer3: not found at this Tm N.d. 0.00
mm4__chr4 53.48 Mbp_chr4_53477346_R Primer3: not found at this Tm N.d. 0.00
mm4__chr2 65.33 Mbp_chr2_65334643_F TCGTCGGCAGCGTCTGCTTTCCTTACTCCCCTGT 58.2 0.00
mm4__chr2 65.33 Mbp_chr2_65334643_R GTCTCGTGGGCTCGGCACCAAAGCCAATCCAACGT 59.3 0.00
mm4__chr18 7.60 Mbp_chr18_7600890_F TCGTCGGCAGCGTCAGGTGCTGTCAGGGTTTCAG 59.8 0.00
mm4__chr18 7.60 Mbp_chr18_7600890_R GTCTCGTGGGCTCGGGGTGAGCCTGTTCTCAGTCC 60.0 0.00
mm4__chr17 21.70 Mbp_chr17_21702869_F TCGTCGGCAGCGTCCAACCCCTACAGCATCGTGT 60.0 0.00
mm4__chr17 21.70 Mbp_chr17_21702869_R GTCTCGTGGGCTCGGCTTCTTGACGAAGCGGTTGC 60.1 0.00
mm4__chr17 21.42 Mbp_chr17_21415425_F TCGTCGGCAGCGTCCATCGTGTCATCGGAGGAGG 59.9 0.00
mm4__chr17 21.42 Mbp_chr17_21415425_R GTCTCGTGGGCTCGGCTTCTTGACGAAGCGGTTGC 60.1 0.00
mm4__chr11 61.84 Mbp_chr11_61838394_F TCGTCGGCAGCGTCGAGGTCTCATGGGCTGTTGT 59.6 0.00
mm4__chr11 61.84 Mbp_chr11_61838394_R GTCTCGTGGGCTCGGGGTACCACTCCTCTCCTTGT 58.0 0.00
mm1__chr20 38.62 Mbp_chr20_38620718_F TCGTCGGCAGCGTCTGGGACTTGGTCTCCAGTGA 60.1 0.00
mm1__chr20 38.62 Mbp_chr20_38620718_R GTCTCGTGGGCTCGGAGAGCCTGAAGGAAGGGACA 60.1 0.00

Off-target amplicon sequences

Primers underlined, off-targets in bold.

ontarget_mm0__chr7 5.53 Mbp_chr7_5529602 CACGATGGAGGGGAAGACGGCCCGGGGGGCATCGTCGCCCGCGAAGCCGGCCTTGCACATGCCGGAGCCGTTGTCGACGA
CGAGCGCGGCGATATCATCATCCATGGTGAGCTGCGAGAATAGC
mm0__chr5 77.78 Mbp_chr5_77784918 ATATCACCGCGCTCGTCATTGACAACGGCTCCGGCATGTGCAAGGCCAGCTTCACGGGCGACAATGCCGCCCGGGCAGTC
TTCCCCTCCATCGTT
mm1__chr22 15.72 Mbp_chr22_15721518 ATACTGCCGTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCAGGTGACGATGCCCCCCGGGCTGTC
TTCCCTTCCATCGTGGGGTGCCCCAGGCACCAGAACATGAT
mm1__chr2 131.63 Mbp_chr2_131626170 ATACCGCTGTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCGGGCGACGATGCCCCCCAGGCTGTC
TTCCCTTCTATCGTGGGGCGCCCCAGGCACCAGGGCATGATGGAGGGCATGCATCAGAAG
mm1__chr2 131.26 Mbp_chr2_131263587 TGGATGATGATACCGCCGTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCGGGCGACGATGCCCCC
CGGGCTGTCTTCCCTTCCATC
mm1__chr2 130.66 Mbp_chr2_130656781 ATACCGCTGTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCGGGCGACGATGCCCCCCGGGCTGTC
TTCCCTTCCATC
mm1__chr2 130.46 Mbp_chr2_130463888 GATGGAAGGGAAGACAGCCCGGGGGGCATCGTCGCCCGCAAAGCCGGCCTTGCACATGCCAGAGCCGTTGTCAATGACGA
GCACGGCGGTATCATCAT
mm1__chr2 130.08 Mbp_chr2_130075316 GATGGAAGGGAAGACAGCCCGGGGGGCATCGTCGCCCGCAAAGCCGGCCTTGCACATGCCAGAGCCGTTGTCAATGACGA
GCACAGCGGTAT
mm1__chr14 19.40 Mbp_chr14_19402853 ATCATGTTCTGGTGCCTGGGGCACCCCACGATGGAAGGGAAGACAGCCCGGGGGGCATCGTCACCTGCAAAGCCGGCCTT
GCACATGCCAGAGCCGTTG
TCAATGACGAGCACGGCAGTAT
mm1__chr14 19.00 Mbp_chr14_18998899 ATACTGCCGTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCAGGTGACGATGCCCCCCGGGCTGTC
TTCCCTTCCATCGTGGGGTGCCCCAGGCACCAGAACATGAT
mm1__chr9 100.73 Mbp_chr9_100731271 GCCAGCTCACCATGGATGATGATATCGCCCCACTTGTCATTGACAACGGCTCAGGCATGTGCAAGGCCGGCTTTGCAGGT
GATAATGTCCCCCTGGGCCATCTTCCCCCTCCATCGTGGAGCTCCCTAG
mm1__chr1 223.86 Mbp_chr1_223864796 CATGGAAGGGAAGATGGCCCAGGGGTCATCATCCCCTGTGAAGCCAGCCTTGCACATGCCGGAGCTGTTGTCAATGACGA
GCGCCGCGATATCATCATCCATGGTGAGCTGGC
mm2__chr22 41.08 Mbp_chr22_41075184 ATGCCCACCATCATGACCTGGTGCCTGGTGCACCCCACGATGGAAGGGAAGATGGCCCAGGGGACATCGTCACCCGCGAA
GCCGGCCTTGCACATGCCAGAGCTGTTGTCAACAACGAGCGCAGCAATA
mm1__chr19 9.52 Mbp_chr19_9518181 GCTGTCAGCTCACCATGGATGATGATATTGAGGTGCTTGTAGTTAACAATGGCTCCGGCATGTGCAAGGTCAACTTCGTG
GGCGACAATGCCCCCCGGGCCATCTTCCCCTCCATCGTGG
mm2__chrX 46.29 Mbp_chrX_46289073 TGGAGGGGAAGACAACTGGGGGGCATCATCGCCCATGAAGGCAGCCTTGCACATGCCAGAGCCTTTGTTGAAGACGAGAG
CAGTGATATCATCATTCATGGTGATCTGGCAGTGGGTGTGGACTGGTGATGGAG
mm3__chr5 80.30 Mbp_chr5_80302792 ATGATGAAGGGGAAGGCAGCCCGGGGGACACCATCGCTCGCAAAGCTAGCCTTGCATATGCCAGAGCCATTGTTGAATAC
GAGAGCAGTGATATCGTCATGCATGGTGAGCT
mm2__chr15 43.99 Mbp_chr15_43990129 CATCGTCACCTGCAAAGCTGGCCTTGCACATGCCAGAGCCATTGTCGACGACAAGCATGGTGAT
mm3__chr11 1.80 Mbp_chr11_1804040 CACGATGGAGGGGAAGACGGCCCAGGGGGCATCGTCGTCGGTGAAGCTGGCCTTGCACATGCCAGAGGTGTTGTCTATGA
CAAGCAAGGTGATATTATCATCCATGGTGAGCTGGG
mm2__chr5 98.34 Mbp_chr5_98338775 GCAGCTAGCTCACCATGGATGATGATATCAGCACGCTCCTCATCAACAAGGGCTCTGGCATGTGCAAGGCCGGCTTCACA
GGTGACAATGC
mm4__chr8 35.75 Mbp_chr8_35745491 CCAAGGAGCCTTAAATGCCCACCATGCACATGCCTAAGGCATTGAGAATTTAAGTCAAAGCGTACACTGATGCAATGACA
mm4__chr1 2.17 Mbp_chr1_2172168 GGACGGGCACATCAAGCTCACAGACTACGGCATGTGCAAGGTGCGTGCCTTGGACCGCCTCCCCTGACCATCCCGCATGT
GCGTCTCGGGGCGCCTGTCCCGCGGGGTAGTGTCTACAAG
mm4__chr12 27.78 Mbp_chr12_27780428 GAGCAGAGCGACTACTTCCGCGCCCTCTACCGCTCCGGCATGCGCGAGGCCCTGAGCCAGGAGGCCGGCGGCCCGGAGGT
GCAGCAGCTGCGCGGCCTCAGCGCGCCGGGCCTGCGGCTGGTGCTGGACTTCATCAACGC
mm3__chr6 88.28 Mbp_chr6_88276931 CCCACAATGGAGGGGAAGACGGCCTGGGGAGCCTTGCGCATGCTGGAGCAGTTGTCGACGACGACGAGCGCGGTGATAGC
ATCATCCATGGTGAGCTGGC
mm2__chr5 61.83 Mbp_chr5_61832990 GACGTTGTGTCCAGCAAAGCCAGCTTTGCACATGCTGGAGCCATTGTCAATGACCAGCCCATGATCTCTTCTTTCACTGC
TTTGGCCAAGGA
mm2__chr20 1.16 Mbp_chr20_1160615 GCTCCTCCACCGATCACAATGGAAGAAGAGATCACCATACTCGTCATTGACAATGGCTCCAGCATGTGCAAAGCTGGTTT
TGCTGGGGACGACGCCCCTGGAACCATGTTTCCC
mm2__chr1 237.93 Mbp_chr1_237926862 CTCCTCCACCAGTCACGATGGAAGAAGAGATCGCCACGCTTGCCATTGACAATGGCTCCAGCATGTGCAAAGCTGGCTTT
GCTGGGGACGATGCCCCCCAAGCTGTGTTTCCCTCCA
mm1__chr17 81.51 Mbp_chr17_81512299 CCATCCACTCACCTGGTGTCTGGGGCGCCCGACGATGGAAGGAAACACGGCTCGGGGAGCGTCGTCCCCAGCAAAACCAG
CTTTGCACATGCCGGAGCCATTGTCAATGACCAGCGCGGCGATCTCTTCTTCCAT
mm3__chr8 84.95 Mbp_chr8_84948621 CTGCTTTGCCACTGGTTCACATCTGCCCTGAGCTCACCATGGATGATGATACCTCTGCACTTGTTGGCAACTGCTCTGGT
ATGTGCAAGG
CCAGAATCCTTGTCAAAGATGCCCCCAGGCCTTCTTCCTTTC
mm2__chrX 53.14 Mbp_chrX_53143858 GATGGAGGGGAACACAGCTCAGGGGCCATCCATCCCCAGCAAAGCCAGCTTTGCACATGCCAGAGCCATTGTCAATGATG
AGCGCGGCGATCTCTATTTCCATTGCAACCTGTGGAAGAGCA
mm4__chr5 138.28 Mbp_chr5_138283423 TAGGGCAAGAGCATGTGCAAACCTTGTCAAATAAAAAAATTAAAAGCCAGTCCAACCACTCAGGCACGTGCATGGGCCGC
CCTATGGCTACAACCCCCCTTCCCTCCCACTCACTTT
mm4__chr3 180.82 Mbp_chr3_180820575 AATGATATCGCCACGCTCGTCATGGACAATGGCTCTCCCATGTGCAAGGCCAGCTTAGCAGGCGACGATGCCCCTCCATC
GTGAGGCACCCATGGCACCAGGGCATGATCGTGGGCATGGGTCAGAAGAA
mm4__chr3 49.17 Mbp_chr3_49171474 CCAGTGTCCAAGTGTCCGTTTATCTGTCCGTTTGCATATGACCGTAGATCGGCACGCCCTGCTCCAGCTGCGGCATGTGC
AGGG
TTAAGGCGAGAGGAGGTTTCC
mm3__chr2 73.90 Mbp_chr2_73901346 GTGTGAAGAGGAGACCACCGCGCTCGTGTGTGACAATGGCTCTGGCCTGTGCAAGGCAGGCTTCGCAGGAGATGATGCCC
CCCGGGCTGTCTTCCCCTCCATTGTGGG
mm4__chr3 59.58 Mbp_chr3_59579128 TGCACTGATGTGAAGCCCATCTGGGATCTCTAGACACAAGGCCACCGGCATGAGCATGGCAGCCCAGGGAGAATACAGAT
GGTGTACCTCTATGCCAAACGGATCAGAATGCT
mm4__chr12 132.07 Mbp_chr12_132067382 CAGGACAGACCGTGGTGGCCCAGCCCGTGCACATNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN
GCCATCCAGCCCCAGGCTGCACAGGGCCCGGCAGCCGTCCAGCAGAAGGTACCGG
mm4__chr2 67.82 Mbp_chr2_67823888 TTTGGCAGCCTCATGGTTCAGCAATACCAGTGGTCCCTCACCTTTGGCTTCGGGCGCCGTGCACAAGCCGGCTCCGGTGA
GCCACCAGCCATCACCCACAGTGCTCTT
mm1__chr9 6.83 Mbp_chr9_6834487 CCGCTACCAGCTCACCATAGATGATGATATTGCTACGCCCGTCGTCGACAACGGCTCCGGCATGTGGAAGGCTGGCTTCG
CAGGTGACGATGACCCCCGGCCGTCTTCCCCTTCATCGTGT
mm4__chr2 57.76 Mbp_chr2_57764984 AGATTGCTGGCTCGTCAATGATAGTGGCTCTGGCATGTGCAAAGCTGGCTTTGCTGGGGAAGACACCCCCTGAGCCATGT
TTCTCTCCATCGTCAGGTGCCCCTGGCACCACAGTGTGATAGTGGGCATGGGCAAGAAG
mm4__chr2 211.78 Mbp_chr2_211778667 GGAACCGGCTGGACAGTTTATCCCCCTTTAGCAGGAAACCTAGCACATGCAGGAGCCTCTGTGAACCTGACCATCTTCTC
GCTCC
mm4__chr2 202.62 Mbp_chr2_202619468 GGAGCGAGAAGATGGTCAGGTCCACAGAGGCTCCTGCATGTGCTAGGTTTCCTCTTAAAGGGGGATAAACCGTCCAGCCG
GTTCCAGCGCCGGCTTCTACTATTGA
mm4__chr1 106.81 Mbp_chr1_106805614 CACCTGCTAAGTGGAGCGAGAAGATGATCAGATCCAGAGGCTCCTGCATGTGCTAGGTTTCCTGCTAAAGGGGGATAAAC
TGTTTAGCTGATTCAAGCGCCAGC
mm4__chr13 111.35 Mbp_chr13_111346406 ATCAGGGACAGCCAGCCATGGCCCGGATGTGTGACAACCCTGCACAGGCCGGGGCCTCTGCCCTCGCGCAGTTGGTGCCC
TGCTGAGCGAGGCGAAGGCCAGACAGAGCTGCCAAGGAA
mm3__chr9 135.73 Mbp_chr9_135728729 GCCCAGCTTGGAGAGACATCTCCCCAGCGGCCCCGGCATGTGCTCAGCAGCTGGTGAGCTCAATCCAGGAGTGTTTTGCC
GGGTCTGGTTA
mm3__chr5 131.66 Mbp_chr5_131660060 ATGCCCATCATCACACCCTGGTGCCTCGGGTGCCCCATTATGGAGGGGAAGACGGCCCGGGGGCATCGTTGCCCGTGAAG
CAGACCTTCCACATGCCAGAGCCATTGTCGACGACAAGCACAGCAAT
mm2__chr6 46.21 Mbp_chr6_46205009 ATGGAAGAAGAGATCCCCGCGCTCTTCATTGACAATGGCTCCGGCATGTGAAAAGCAGCTTTGCTGGGACAATGCCCTCC
GAGCCATAT
mm4__chr5 135.07 Mbp_chr5_135066866 AGGGAGGCTGATGTGAAAGCCAGCATCCACCGGCCTCGGCATGTGCCAGGCTCTGTGCTTTCCATACACCAACCCCAGGA
mm4__chr17 16.47 Mbp_chr17_16466367 CTTTCAGACTCGGCACCCATGGCCTGCCCTCCTTCTCAGGAAGACACAGAATGGGGGTAGGCACACAAGGCCTGCTGCAT
GTGCAAGG
GAAGTGGAAGAATCAGATCCAGGTGTGGTCCCTG
mm3__chrY 17.76 Mbp_chrY_17758070 AACGGTGGAGGGAAACATGGCTTGGGGAGCATCGTCCCCTGCAAAGCCAGCTTTGCACATGCAAGAGCCATTGTCAATGA
CGAGCGCAGCGATCTCTTCTTCTACTGCGATGGACAGAGGAGTGGGACAGAGGA
mm4__chr6 169.65 Mbp_chr6_169648288 TCTACAGATGCCTTCCGTGCACATGCCAATGCCTTCCGTGCACACGCCCGAGCCCTGGGTGGCATTTGGGGCCTCGCCAC
CACTGCTACCCAGCAGCTCCCAGCAGAAGGCCCAAGAGAGCCAAATGCAGAAGAGCCAG
mm4__chr3 46.88 Mbp_chr3_46875620 CACCACCTCCCTGGATTTCCCAACTGCTCCACAATGTGCATGGGGCAAGAACTGTTACGACGCTGCA
mm4__chr10 5.59 Mbp_chr10_5585443 TTCTGAGGACTCACAGCTGCTGAGGAAACACACTTCCACTACCAGCTTCACACCCAGCACGTGCGGGAGCCGTGGAAAGC
CACCACCCTTGTCTCCCTTC
mm4__chr19 31.57 Mbp_chr19_31566196 TTCACCAGAGCCCAACGTACAGCGGCAGCTGGCAGAGAAGTGAGTGCGGGAGCACTTGATTGAATCTTCATGTAGCCAGG
CCCTCCACATGCCTGAGCCACTGTACTGGGGCATAGGGTAGGG
mm4__chr20 59.74 Mbp_chr20_59737337 TCTGTGGGGATCCTGGTTCAGGGCTCTCTATGTCCCCCTGGCACGTGCCTGAGCTGTTGCTGGCACTGGAGATACAGCTG
CA
mm4__chr1 248.18 Mbp_chr1_248176264 CAGTGACTTCTCAATTTCCGACCATGATTTTCATGGTTTCTGTGTTACTATTGCTCTGGCATGTGCACAGCACCACACTG
TCATGAAATCTGA
mm4__chrX 152.91 Mbp_chrX_152906792 GTCTGCTCAGCTCCTGGATGCAAAGCATCCCAATTCACCTTTGGGAAAGAAGCCCTTCTGCTTGCACCTCTGCCTGTGGC
ACACCTGCTCCTGCTTGTGCATGGCGCCCAGTGCCCAGCACACATGCACTGAGACAAGCC
mm4__chr3 74.00 Mbp_chr3_74002249 AGAGCAGCCATGAACTGACAACTCCCAGTAAAGCAGACACTCTGCACATGCCAGTGCAGGTGGGTGGCACCAACACTTTG
CTCAA
mm4__chr2 205.39 Mbp_chr2_205387855 TGGGTAATGGTTGCCAGTCCAGCCGTCATCTGAGTGACGCTGCTGGTGTTCCCTGCACCTTCCAGAGCCGTTCAGAAACA
CAGGCTGCCAGTT
mm4__chr7 157.09 Mbp_chr7_157087829 TGGCATACAGTAGGGCTTGCTATTGACGATGAGCGCACCCACGGCTCCGGCAACTACAAGGCGGTGCTCACGGACAGCAC
GGCCCTCGGCGTGGATATCCGGAAGGCAG
mm4__chr2 65.33 Mbp_chr2_65334643 TGCTTTCCTTACTCCCCTGTCAAAGGCTCGGGCATCAGCAGGGCTTTGGAAAGTAAGTCCAAACTTCCTATTATCGACCT
TCCAGTGATGAAACGTTGGATTGGCTTTGGTG
mm4__chr18 7.60 Mbp_chr18_7600890 AGGTGCTGTCAGGGTTTCAGAGGAAACTGCTCCTGCATCTGCAGGGAGCTCTGCTGGACTGAGAACAGGCTCACC
mm4__chr17 21.70 Mbp_chr17_21702869 CAACCCCTACAGCATCGTGTCATTGGAGGAGGACGGGCTGCACCTGGTCACCATGTCGGGNNNNNNNNNNNNNNNNNNNN
NNNNNN
NNNNNNNNCGGCGCAGGTGCCGCAACCGCTTCGTCAAGAAG
mm4__chr17 21.42 Mbp_chr17_21415425 CATCGTGTCATCGGAGGAGGACGGGCTGCACCTGGTCACCATGTCGGGNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN
NNCGGCGCAGGTGCCGCAACCGCTTCGTCAAGAAG
mm4__chr11 61.84 Mbp_chr11_61838394 GAGGTCTCATGGGCTGTTGTTTTGTTCAACAATGAAGTCTCCATGTAAATGGCGGAGCCGTTTATAGAAGAAAGTTNNNN
NNNNNNNNNNNNNNNNNNNNNNNNNNNNNGGAGAGTTGTTACAAGGAGAGGAGTGGTACC
mm1__chr20 38.62 Mbp_chr20_38620718 TGGGACTTGGTCTCCAGTGAAGCTTCTGCTCTGTGCAAATGCCGGAGCCGTTGCCAGGAGGTTGTGCCCTGTCCCTTCCT
TCAGGCTCT

Input file for Crispresso

Crispresso, written by Luca Pinello, is a software package to quantify the Cas9-induced mutations on off- or on-targets.

Click here to download an amplicon input file for Crispresso. For each off-target, it includes the off-target name, its PCR amplicon and the guide sequence. Keep a copy of this file.

After sequencing, run CRISPRessoPooled. The tool will map the reads to the amplicons and analyse the mutations:
CRISPRessoPooled -r1 Reads1.fastq.gz -r2 Reads2.fastq.gz -f crisporAmplicons_gYvicTzp9e5VPFC9YwLR.txt --name MY_EXPERIMENT

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CRISPR/Cas9 Guide Designer for chordate vertebrate ecdysozoans lophotrochozoans protostomes spongi corals plants butterflies metazoans genomes fruitflies insects nematodes mammals.