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PCR primers for off-targets of AGGACTCGGCGCGCCGGAAG TGG

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_5529712_F TCGTCGGCAGCGTCTGGTGAGCTGCGAGAATAGC 60.1 1.00
ontarget_mm0__chr7 5.53 Mbp_chr7_5529712_R GTCTCGTGGGCTCGGCGGCTTCCTTTGTCCCCAAT 60.6 1.00
mm4__chr1 167.49 Mbp_chr1_167492268_F TCGTCGGCAGCGTCGCGTGCTCACAGTTGACATG 60.1 0.42
mm4__chr1 167.49 Mbp_chr1_167492268_R GTCTCGTGGGCTCGGGTCTCCTGGTCTAGTTGGCG 59.8 0.42
mm4__chr2 36.67 Mbp_chr2_36671112_F TCGTCGGCAGCGTCACCACACTTCTGTAAGATGAGTC 57.8 0.29
mm4__chr2 36.67 Mbp_chr2_36671112_R GTCTCGTGGGCTCGGTCTGTCCTCATTTTGGGTGCA 59.8 0.29
mm4__chr6 30.07 Mbp_chr6_30067205_F TCGTCGGCAGCGTCGAACTAATCCAGCGACGCCT 60.1 0.20
mm4__chr6 30.07 Mbp_chr6_30067205_R GTCTCGTGGGCTCGGCCCTGGGGATAACCTGGCTA 60.4 0.20
mm4__chr17 82.02 Mbp_chr17_82023286_F TCGTCGGCAGCGTCCTTTGTTCTCATTGGCCCGC 60.1 0.19
mm4__chr17 82.02 Mbp_chr17_82023286_R GTCTCGTGGGCTCGGGGGGCCTCAGTATCCAGGA 60.0 0.19
mm4__chr8 3.59 Mbp_chr8_3585073_F TCGTCGGCAGCGTCACAACTCGCTATCAGCTGGG 59.8 0.19
mm4__chr8 3.59 Mbp_chr8_3585073_R GTCTCGTGGGCTCGGACCACTAGGGGTCAGTCACA 59.8 0.19
mm4__chrX 149.53 Mbp_chrX_149526013_F Primer3: not found at this Tm N.d. 0.16
mm4__chrX 149.53 Mbp_chrX_149526013_R Primer3: not found at this Tm N.d. 0.16
mm4__chr1 7.77 Mbp_chr1_7771217_F Primer3: not found at this Tm N.d. 0.13
mm4__chr1 7.77 Mbp_chr1_7771217_R Primer3: not found at this Tm N.d. 0.13
mm4__chr9 37.80 Mbp_chr9_37800772_F TCGTCGGCAGCGTCGTGTCTCAGCTTTCCCGGTC 60.6 0.12
mm4__chr9 37.80 Mbp_chr9_37800772_R GTCTCGTGGGCTCGGCCTCAGTGCCCCTGTTCAG 60.0 0.12
mm4__chr7 2.69 Mbp_chr7_2688085_F TCGTCGGCAGCGTCTTTGGCTTTAGGCTCTGGGG 59.9 0.12
mm4__chr7 2.69 Mbp_chr7_2688085_R GTCTCGTGGGCTCGGCAGGACAGGAGGGCCACA 60.9 0.12
mm4__chr3 11.02 Mbp_chr3_11023589_F TCGTCGGCAGCGTCGGGAACTTCTGGACCCACTG 59.9 0.10
mm4__chr3 11.02 Mbp_chr3_11023589_R GTCTCGTGGGCTCGGTTAGACTGGGAGATACTTGTGC 57.2 0.10
mm3__chr19 50.15 Mbp_chr19_50147962_F TCGTCGGCAGCGTCCTTCCTAGGGATCGCAGCAG 59.9 0.10
mm3__chr19 50.15 Mbp_chr19_50147962_R GTCTCGTGGGCTCGGTCATTCCCTGAGTCCCACCT 59.8 0.10
mm3__chr5 74.14 Mbp_chr5_74143932_F Primer3: not found at this Tm N.d. 0.10
mm3__chr5 74.14 Mbp_chr5_74143932_R Primer3: not found at this Tm N.d. 0.10
mm4__chr4 48.34 Mbp_chr4_48341908_F Primer3: not found at this Tm N.d. 0.10
mm4__chr4 48.34 Mbp_chr4_48341908_R Primer3: not found at this Tm N.d. 0.10
mm4__chr19 49.12 Mbp_chr19_49119873_F TCGTCGGCAGCGTCTCCCGGATTCCCTTCCTCAT 60.0 0.09
mm4__chr19 49.12 Mbp_chr19_49119873_R GTCTCGTGGGCTCGGCGGGATTCTAGGGTTCGGTC 59.6 0.09
mm4__chr10 81.87 Mbp_chr10_81874994_F TCGTCGGCAGCGTCGAGGATGTCGGCGCTCCTG 62.7 0.07
mm4__chr10 81.87 Mbp_chr10_81874994_R GTCTCGTGGGCTCGGCCTCCGGTCGGTCAGAGA 60.0 0.07
mm4__chrX 130.06 Mbp_chrX_130060695_F TCGTCGGCAGCGTCCCATCGCTGGCTCGACAG 60.5 0.06
mm4__chrX 130.06 Mbp_chrX_130060695_R GTCTCGTGGGCTCGGTGAGTGAGCTCCTTCCCCTC 60.6 0.06
mm4__chr1 154.57 Mbp_chr1_154567762_F TCGTCGGCAGCGTCCCTACCACGGCAGAACCAAT 60.0 0.05
mm4__chr1 154.57 Mbp_chr1_154567762_R GTCTCGTGGGCTCGGGGCTGCTCTTCCAGGGAATA 59.1 0.05
mm4__chr17 45.78 Mbp_chr17_45783357_F TCGTCGGCAGCGTCAGCCAATTCAGAAGACGGCC 60.6 0.04
mm4__chr17 45.78 Mbp_chr17_45783357_R GTCTCGTGGGCTCGGGAACGCTGCTAGGAGCCC 60.2 0.04
mm4__chr15 77.63 Mbp_chr15_77631250_F Primer3: not found at this Tm N.d. 0.04
mm4__chr15 77.63 Mbp_chr15_77631250_R Primer3: not found at this Tm N.d. 0.04
mm4__chr2 11.93 Mbp_chr2_11925893_F Primer3: not found at this Tm N.d. 0.04
mm4__chr2 11.93 Mbp_chr2_11925893_R Primer3: not found at this Tm N.d. 0.04
mm4__chr9 93.92 Mbp_chr9_93921356_F TCGTCGGCAGCGTCCCTCTGCGTTTCCAAGTTGC 60.0 0.04
mm4__chr9 93.92 Mbp_chr9_93921356_R GTCTCGTGGGCTCGGGCACTGGGTGTGGTTTTGTG 60.1 0.04
mm4__chr9 137.59 Mbp_chr9_137590329_F Primer3: not found at this Tm N.d. 0.03
mm4__chr9 137.59 Mbp_chr9_137590329_R Primer3: not found at this Tm N.d. 0.03
mm4__chr11 1.04 Mbp_chr11_1037090_F TCGTCGGCAGCGTCCGGGATGGACCTGTTGGG 59.7 0.02
mm4__chr11 1.04 Mbp_chr11_1037090_R GTCTCGTGGGCTCGGATTCAGGTGCAGCAGGGTTC 60.6 0.02
mm4__chr14 32.93 Mbp_chr14_32933398_F Primer3: not found at this Tm N.d. 0.02
mm4__chr14 32.93 Mbp_chr14_32933398_R Primer3: not found at this Tm N.d. 0.02
mm3__chr14 56.12 Mbp_chr14_56119059_F TCGTCGGCAGCGTCGAAGTGGCCAGAACTCGGG 60.3 0.02
mm3__chr14 56.12 Mbp_chr14_56119059_R GTCTCGTGGGCTCGGCACATGGAGTCCCTGCCC 59.7 0.02
mm4__chrX 9.47 Mbp_chrX_9465457_F TCGTCGGCAGCGTCCACAAGTTGCGCTGCTCG 60.1 0.02
mm4__chrX 9.47 Mbp_chrX_9465457_R GTCTCGTGGGCTCGGGCCTCCCCGCAGAAGTTT 59.9 0.02
mm4__chr2 239.35 Mbp_chr2_239351879_F TCGTCGGCAGCGTCTGTGGACGCAAGACAGAAGT 59.5 0.01
mm4__chr2 239.35 Mbp_chr2_239351879_R GTCTCGTGGGCTCGGCAGAGCCAGTCCTGTGCTG 60.3 0.01
mm4__chr1 6.26 Mbp_chr1_6260748_F TCGTCGGCAGCGTCCGATAGGGCGCGGAAAGC 61.3 0.01
mm4__chr1 6.26 Mbp_chr1_6260748_R GTCTCGTGGGCTCGGCGCTCGAATGCAGGGTGA 60.1 0.01
mm4__chr15 42.02 Mbp_chr15_42024149_F TCGTCGGCAGCGTCTAAGTGGGATGGGAGGGGAG 60.0 0.01
mm4__chr15 42.02 Mbp_chr15_42024149_R GTCTCGTGGGCTCGGGGGGATGTCACTGTTTGCCT 60.2 0.01
mm4__chr1 157.99 Mbp_chr1_157992657_F TCGTCGGCAGCGTCGGGATGGGTGGAAGGCTAAG 59.8 0.01
mm4__chr1 157.99 Mbp_chr1_157992657_R GTCTCGTGGGCTCGGCTCTTGGACCTGCCTTGAGG 60.0 0.01
mm4__chr18 61.33 Mbp_chr18_61334100_F TCGTCGGCAGCGTCCTCTCTGGAAGCAGGGAAGC 60.1 0.01
mm4__chr18 61.33 Mbp_chr18_61334100_R GTCTCGTGGGCTCGGTGGATCCCCAGTCACCATCC 60.9 0.01
mm4__chr20 56.63 Mbp_chr20_56630187_F Primer3: not found at this Tm N.d. 0.01
mm4__chr20 56.63 Mbp_chr20_56630187_R Primer3: not found at this Tm N.d. 0.01
mm4__chrX 14.53 Mbp_chrX_14529596_F TCGTCGGCAGCGTCCCAGCACAGGATCTCAGCAA 60.0 0.01
mm4__chrX 14.53 Mbp_chrX_14529596_R GTCTCGTGGGCTCGGTCTTTTCTGTTTAGCAAAGCCCT 58.7 0.01
mm3__chr18 72.87 Mbp_chr18_72867651_F Primer3: not found at this Tm N.d. 0.01
mm3__chr18 72.87 Mbp_chr18_72867651_R Primer3: not found at this Tm N.d. 0.01
mm4__chr17 81.44 Mbp_chr17_81442566_F TCGTCGGCAGCGTCTTCGTGCCTTCTGTGGGAC 59.9 0.00
mm4__chr17 81.44 Mbp_chr17_81442566_R GTCTCGTGGGCTCGGGAGACTCCATAGGGTGCAGC 59.8 0.00
mm4__chr11 31.99 Mbp_chr11_31987512_F Primer3: not found at this Tm N.d. 0.00
mm4__chr11 31.99 Mbp_chr11_31987512_R Primer3: not found at this Tm N.d. 0.00
mm4__chr6 31.90 Mbp_chr6_31902066_F Primer3: not found at this Tm N.d. 0.00
mm4__chr6 31.90 Mbp_chr6_31902066_R Primer3: not found at this Tm N.d. 0.00
mm4__chr6 158.56 Mbp_chr6_158560014_F TCGTCGGCAGCGTCCTCAGGGCTAGCCTGGGT 60.3 0.00
mm4__chr6 158.56 Mbp_chr6_158560014_R GTCTCGTGGGCTCGGCGGAAGCGACTCTCACGG 60.2 0.00
mm4__chr3 138.07 Mbp_chr3_138068633_F TCGTCGGCAGCGTCCTCCAGGGTCGTGTTGTCAT 59.6 0.00
mm4__chr3 138.07 Mbp_chr3_138068633_R GTCTCGTGGGCTCGGGGTAGCAGCCTTGGTGAAGT 59.9 0.00
mm4__chr2 232.49 Mbp_chr2_232487829_F Primer3: not found at this Tm N.d. 0.00
mm4__chr2 232.49 Mbp_chr2_232487829_R Primer3: not found at this Tm N.d. 0.00
mm4__chr19 42.70 Mbp_chr19_42698514_F Primer3: not found at this Tm N.d. 0.00
mm4__chr19 42.70 Mbp_chr19_42698514_R Primer3: not found at this Tm N.d. 0.00
mm4__chr17 44.54 Mbp_chr17_44539941_F Primer3: not found at this Tm N.d. 0.00
mm4__chr17 44.54 Mbp_chr17_44539941_R Primer3: not found at this Tm N.d. 0.00
mm4__chr16 58.03 Mbp_chr16_58026809_F TCGTCGGCAGCGTCTACATTCAGCCTGTGTCCCG 59.7 0.00
mm4__chr16 58.03 Mbp_chr16_58026809_R GTCTCGTGGGCTCGGGAGGGAAGGAAGGAGGGGAG 60.6 0.00
mm4__chr16 30.33 Mbp_chr16_30334910_F TCGTCGGCAGCGTCGTGGGTGGAGGGCCTAGG 61.0 0.00
mm4__chr16 30.33 Mbp_chr16_30334910_R GTCTCGTGGGCTCGGGGAGCTGGAATGACTGGCAA 60.3 0.00
mm4__chr16 29.59 Mbp_chr16_29594822_F TCGTCGGCAGCGTCGTGGGTGGAGGGCCTAGG 61.0 0.00
mm4__chr16 29.59 Mbp_chr16_29594822_R GTCTCGTGGGCTCGGGGAGCTGGAATGACTGGCAA 60.3 0.00
mm4__chr16 22.44 Mbp_chr16_22437533_F TCGTCGGCAGCGTCCTGGAATGACTGGCAACCCA 60.2 0.00
mm4__chr16 22.44 Mbp_chr16_22437533_R GTCTCGTGGGCTCGGGGAGGAGACCCAGGAAGCT 60.3 0.00
mm4__chr16 18.93 Mbp_chr16_18925885_F TCGTCGGCAGCGTCGAGGGGTGGAGGGCCTAG 60.4 0.00
mm4__chr16 18.93 Mbp_chr16_18925885_R GTCTCGTGGGCTCGGCTGGAATGACTGGCAACCCA 60.2 0.00
mm4__chr11 60.92 Mbp_chr11_60924401_F TCGTCGGCAGCGTCCCTCTGTTGCTTCCAGGGAC 60.3 0.00
mm4__chr11 60.92 Mbp_chr11_60924401_R GTCTCGTGGGCTCGGAATCGCAGCACATGGGCT 60.0 0.00
mm4__chr1 43.43 Mbp_chr1_43427702_F TCGTCGGCAGCGTCCAGACAGAGAGTGCGGATGG 60.1 0.00
mm4__chr1 43.43 Mbp_chr1_43427702_R GTCTCGTGGGCTCGGAAGGCACTCACCGGAAGATG 60.0 0.00
mm4__chr1 232.81 Mbp_chr1_232805709_F TCGTCGGCAGCGTCCGCTGTTGGTTTACCCTCCT 59.9 0.00
mm4__chr1 232.81 Mbp_chr1_232805709_R GTCTCGTGGGCTCGGGTGCAGGGTAGCAGGCTG 60.4 0.00
mm3__chr7 35.04 Mbp_chr7_35036979_F TCGTCGGCAGCGTCAAAAGTCGGAGAGGGCTGC 60.0 0.00
mm3__chr7 35.04 Mbp_chr7_35036979_R GTCTCGTGGGCTCGGGGGACGCGGAACTTTGAGG 61.0 0.00
mm3__chr3 51.39 Mbp_chr3_51391273_F Primer3: not found at this Tm N.d. 0.00
mm3__chr3 51.39 Mbp_chr3_51391273_R Primer3: not found at this Tm N.d. 0.00

Off-target amplicon sequences

Primers underlined, off-targets in bold.

ontarget_mm0__chr7 5.53 Mbp_chr7_5529712 TGGTGAGCTGCGAGAATAGCCGGGCGCGCTGTGAGCCGAGGTCGCCCCCGCCCTGGCCACTTCCGGCGCGCCGAGTCCTT
AGGCCGCCAGGGGGCGCCGGCGCGCGCCCAGATTGGGGACAAAGGAAGCCG
mm4__chr1 167.49 Mbp_chr1_167492268 GCGTGCTCACAGTTGACATGTGTGAAGGACCAGGAGAGCCGGAAGAGGTCAGCAGGCTCAGGGGGTGCTGTTAGCCTGTT
TCACCGACAGAAAGGCTTAACCCCTGCGGCGCCAACTAGACCAGGAGAC
mm4__chr2 36.67 Mbp_chr2_36671112 ACCACACTTCTGTAAGATGAGTCAGGACTGGGAGAGCCAGAAGAGGTTTCCAGGAGGAAGTAAGACTTGATTTGCACCCA
AAATGAGGACAGA
mm4__chr6 30.07 Mbp_chr6_30067205 GAACTAATCCAGCGACGCCTGCGCTTTGACGCATTTGGTGCCGTGGAAGGGAAAAAGGGGGACTGCAGTATGCGTCACAC
CCGGAAGCGGCGAGCCGGAAGTGGGGTTAGCCAGGTTATCCCCAGGG
mm4__chr17 82.02 Mbp_chr17_82023286 CTTTGTTCTCATTGGCCCGCACCAAGCGCTGCCTGCGCAGAGCCTGCTAGGAGTTGTAGTTCCCGCGGCCTCTCCCGGCG
CGCCTTGTTCT
TCCTGGATACTGAGGCCCC
mm4__chr8 3.59 Mbp_chr8_3585073 ACAACTCGCTATCAGCTGGGCCATACACCCTCCCTCCCCTTCCGGTGGGGCGGGTCCTGCAAAGCTGTATCCCCTGCATG
TGACTGACCCCTAGTGGT
mm4__chr9 37.80 Mbp_chr9_37800772 GTGTCTCAGCTTTCCCGGTCCCCGGCGGACGGTGGCCGAGGGGCGGCGCGGGTTTCCAGCCGGTGGGGTTAGGCCGCTGC
ACGCCGGAAGTGG
CAGCTGAACAGGGGCACTGAGG
mm4__chr7 2.69 Mbp_chr7_2688085 TTTGGCTTTAGGCTCTGGGGCCCCCCTGCCCGGGTGTTTCTGGGAGGCTGGAGGGTCCCACGTGAGCTCCACCGCGGGCA
GCTGGAGGGAGTGGGGCAGGACGCTGAGCGCCGGGAGGGGTGTGGCCCTCCTGTCCTG
mm4__chr3 11.02 Mbp_chr3_11023589 GGGAACTTCTGGACCCACTGGGTTCTCAAATCCAAGGCTGGTGTAAAAGTATCTCTGATCCATTCCTAGGACTAGGGACG
CCGGAGGAGG
GCACAAGTATCTCCCAGTCTAA
mm3__chr19 50.15 Mbp_chr19_50147962 CTTCCTAGGGATCGCAGCAGTCCCACGCTCCGCCGGACTCCTGGCAAACAGCAGACGCCCAATTCCTGCGCGCCGAGTCG
T
CTGAGCCGCACTGATACTAGGTGGGACTCAGGGAATGA
mm4__chr19 49.12 Mbp_chr19_49119873 TCCCGGATTCCCTTCCTCATCCCAGGGGACCCCCGAGCCCAGATTTCACACCCCAACCCCGGGGACTCAGGCGTGCCCCT
CCCGGCGCCCAGAGTCCC
CGACCGAACCCTAGAATCCCG
mm4__chr10 81.87 Mbp_chr10_81874994 GAGGATGTCGGCGCTCCTGGAGCGCACGCAGGCGGCGGNNNNNNNNNNNNNNNNNGGGCGCGGGGACCCGGCGCGCAGGA
GGCGG
CTTGGAGGGCTGCAGACGCGCCCCGCCGCTCTCTGACCGACCGGAGG
mm4__chrX 130.06 Mbp_chrX_130060695 CCATCGCTGGCTCGACAGCGCCACCTGCTGGACGTCGGCCTCCACGTCCGGCGCGCCGGGGCCCGAAGTTCTCGGAGGCT
GGGACCTCAGTCTGGGAGGGGAAGGAGCTCACTCA
mm4__chr1 154.57 Mbp_chr1_154567762 CCTACCACGGCAGAACCAATCGAAGACTATCCCAGCCGGGGACCCCCCTTTTTTTTCCTGGGACCCAGCGTTTCCGCCTC
CGGGGCGCAGACTCCT
CCCCCTCACCGTCCCAATTGTATTCCCTGGAAGAGCAGCC
mm4__chr17 45.78 Mbp_chr17_45783357 AGCCAATTCAGAAGACGGCCCCGGGGTCTGGCGGGGAGGCCCACCGCTGCCCAGAGGGCTCGGCCGGCCGGGAGGGGGCG
CTGTGTCCTCCCCACGCCCTGCCCGCGGGCTCCTAGCAGCGTTC
mm4__chr9 93.92 Mbp_chr9_93921356 CCTCTGCGTTTCCAAGTTGCATCAGCACCACTAGGACTCCGCCAGCAGGAAGGGGGTGGACCCGAGAGACTTGGCCAGGC
CAAGCCATGACCCCATCATTTTGTCTTAACACAAAACCACACCCAGTGC
mm4__chr11 1.04 Mbp_chr11_1037090 CGGGATGGACCTGTTGGGGGAGGTCGGGGGCCCTGGGGCACCTGCTCTGCCCCTTCCGGGGAGCCCAGTGCTGACCCAGC
TTGGGGGGAACCCTGCTGCACCTGAAT
mm3__chr14 56.12 Mbp_chr14_56119059 GAAGTGGCCAGAACTCGGGCGACCCAGCGGGGGCAGCGCCGGGCACCGGGTCCGCGGCCGTATCGTGGGGGCGGGGGCAG
GGGCTCCACTTCCCGCGCGCCGCGTCCGGGCCGGGGAGGGCGGGCAGGGACTCCATGTG
mm4__chrX 9.47 Mbp_chrX_9465457 CACAAGTTGCGCTGCTCGCGACGAGGTGAGTGCGCAGGGCTCGGCGGGCCGGGCGCGGGGTCCGCAAACTTCTGCGGGGA
GGC
mm4__chr2 239.35 Mbp_chr2_239351879 TGTGGACGCAAGACAGAAGTCGATGGCTAAGTGCCCTTGAAGGCCACGGTGCGCTGGAAGTGGCAGGAAGCTGGGCAGCA
CAGGACTGGCTCTG
mm4__chr1 6.26 Mbp_chr1_6260748 CGATAGGGCGCGGAAAGCCCCCTCCCTTCCCGCCGAGTCCTCCCGCCGGGGTCCTCCCGTCCGTCCGCCCTCGGGCCAGG
GCGGCGACGCTCACCCTGCATTCGAGCG
mm4__chr15 42.02 Mbp_chr15_42024149 TAAGTGGGATGGGAGGGGAGCAGAGCAGAGGACTCGGGGAGTTGGAAGTGGACTGGGACAGTGAGTAAGGAGTGTCAGGC
AAACAGTGACATCCCC
mm4__chr1 157.99 Mbp_chr1_157992657 GGGATGGGTGGAAGGCTAAGATGAACAAAGGAAGGGTCAACTGTGCCTCTTCCAGGCCGCTGAGTCCTCCAACGCCTAGC
TGTGCTCAACCTTGACTGGGTGAAGCCAAGGCAGGGCCTCAAGGCAGGTCCAAGAG
mm4__chr18 61.33 Mbp_chr18_61334100 CTCTCTGGAAGCAGGGAAGCTGGGGCGCACCGGGGCAGGTGGAGGACGTGGCGCGCTGGGAGCGGGGACTGCGCGCGCCT
GGGGATGGTGACTGGGGATCCA
mm4__chrX 14.53 Mbp_chrX_14529596 CCAGCACAGGATCTCAGCAATTTTTCCCTCTTCCCCCCACCCACTCCAGCGCGCAGAGTCCTTTCTTCTCTCTCATTTAC
AACTTCTTTTTAAAAGAAAACATTTTCTAGAAAAAGGGCTTTGCTAAACAGAAAAGA
mm4__chr17 81.44 Mbp_chr17_81442566 TTCGTGCCTTCTGTGGGACACCTGGCCGACAAGGGCCGCCCCTTCCAGGCCGCCGAGGCCTGTGCCGTGGCAGGGGAGGG
CAAGGACCGGCACCTGGAGGGAACCATGGCCCCCGACCACGCTGCACCCTATGGAGTCTC
mm4__chr6 158.56 Mbp_chr6_158560014 CTCAGGGCTAGCCTGGGTGCTCCCGGCCGTGGGGCTCCGCCCCGGCACGGGGCCACGAAGGAGGGCCACNNNNNNNNNNN
NNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNTCTTCCGCCGTGAGAGTCGCTTCCG
mm4__chr3 138.07 Mbp_chr3_138068633 CTCCAGGGTCGTGTTGTCATTACTGAAGACCAGCAGGAGTCCGCGGGCCTGAAGAGGGGCTCAGTAGTCCTAGATGGACT
TCACCAAGGCTGCTACC
mm4__chr16 58.03 Mbp_chr16_58026809 TACATTCAGCCTGTGTCCCGGGAGCCGCTTCCGCCGCCCCCAGCCCTTGCCTCCCCTCCTTCCTTCCCTC
mm4__chr16 30.33 Mbp_chr16_30334910 GTGGGTGGAGGGCCTAGGCCGCGCCTCCCAGCCCCGCGGCCCTAGGCCTCGGCCCGCCCGAGGCGGAGCCCGGGAGGTCG
GGGCGGGGTCCCGGGCCGGTCACCCACCTGGGTTGCCAGTCATTCCAGCTCC
mm4__chr16 29.59 Mbp_chr16_29594822 GTGGGTGGAGGGCCTAGGCCGCGCCTCCCAGCCCCGCGGCCCTAGGCCTCGGCCCGCCCGAGGCGGAGCCCGGGAGGTCG
GGGCGGGGTCCCGGGCCGGTCACCCACCTGGGTTGCCAGTCATTCCAGCTCC
mm4__chr16 22.44 Mbp_chr16_22437533 CTGGAATGACTGGCAACCCAGGTGGGTGACCGGCCCGGGACCCCGCCCCGACCTCCCGGGCTCCGCCTCGGGCGGGCCGA
GGCCT
AGGCCCTCCACCCCCGGGAGCCGGGCGCAGCTTCCTGGGTCTCCTCC
mm4__chr16 18.93 Mbp_chr16_18925885 GAGGGGTGGAGGGCCTAGGCCGCGCCTCCCAGCCCCGCGGCCCTAGGCCTCGGCCCGCCCGAGGCGGAGCCCGGGAGGTC
GGGGCGGGGTCCCGGGCCGGTCACCCACCTGGGTTGCCAGTCATTCCAG
mm4__chr11 60.92 Mbp_chr11_60924401 CCTCTGTTGCTTCCAGGGACGTGGCCCTCTTCCCGACCTCGGTCGCCTGAGCGCGCAGGGATTGGGGATCCGTAGGGCAC
TAGGACCCGGCGGGGCGGAGGGGGCAGCCCATGTGCTGCGATT
mm4__chr1 43.43 Mbp_chr1_43427702 CAGACAGAGAGTGCGGATGGGCCCCGGACCCGGTGTCCTGTCTACATCTACAGCTGTTCACTGGAAGCGCTGAGGGAACA
AATGGTTGGCATGCAGCCCCCTCAGGCGCCCCGAGACCTCATCTTCCGGTGAGTGCCTT
mm4__chr1 232.81 Mbp_chr1_232805709 CGCTGTTGGTTTACCCTCCTGACGGCCCCGGCGCTCCCGCCGCCGAACCGTGGCCCGGTGTCATCCGCTTCGGTCGCGGC
AAGTCCT
GCCTCTTCCGCCTGCAGCCTGCTACCCTGCAC
mm3__chr7 35.04 Mbp_chr7_35036979 AAAAGTCGGAGAGGGCTGCAAACTGGCTGGTCCGGCTGGCGGGGCTCGGCGCGGCGGAAGCNNNNNNNNNNNNNNNNNNN
NNNNNNNNNNNNNNNNNNNNNNNNGCCAGCGCCTCAAAGTTCCGCGTCCC

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.