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PCR primers for off-targets of GGCATCGTCGCCCGCGAAGC CGG

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_5529579_F TCGTCGGCAGCGTCCACGATGGAGGGGAAGACG 59.8 1.00
ontarget_mm0__chr7 5.53 Mbp_chr7_5529579_R GTCTCGTGGGCTCGGGCTATTCTCGCAGCTCACCA 60.1 1.00
mm1__chr2 131.63 Mbp_chr2_131626193_F TCGTCGGCAGCGTCGTGCTCGTCATTGACAACGG 59.8 1.00
mm1__chr2 131.63 Mbp_chr2_131626193_R GTCTCGTGGGCTCGGACTCCTTCTGATGCATGCCC 60.1 1.00
mm1__chr2 131.26 Mbp_chr2_131263610_F TCGTCGGCAGCGTCTGGATGATGATACCGCCGTG 59.9 1.00
mm1__chr2 131.26 Mbp_chr2_131263610_R GTCTCGTGGGCTCGGGATGGAAGGGAAGACAGCCC 60.1 1.00
mm1__chr2 130.66 Mbp_chr2_130656804_F TCGTCGGCAGCGTCGTGCTCGTCATTGACAACGG 59.8 1.00
mm1__chr2 130.66 Mbp_chr2_130656804_R GTCTCGTGGGCTCGGGATGGAAGGGAAGACAGCCC 60.1 1.00
mm1__chr2 130.46 Mbp_chr2_130463865_F TCGTCGGCAGCGTCGATGGAAGGGAAGACAGCCC 60.1 1.00
mm1__chr2 130.46 Mbp_chr2_130463865_R GTCTCGTGGGCTCGGGTGCTCGTCATTGACAACGG 59.8 1.00
mm1__chr2 130.08 Mbp_chr2_130075293_F TCGTCGGCAGCGTCGATGGAAGGGAAGACAGCCC 60.1 1.00
mm1__chr2 130.08 Mbp_chr2_130075293_R GTCTCGTGGGCTCGGGTGCTCGTCATTGACAACGG 59.8 1.00
mm2__chr22 41.08 Mbp_chr22_41075161_F TCGTCGGCAGCGTCGGAGTCCTTCTGACCCATGC 60.1 0.79
mm2__chr22 41.08 Mbp_chr22_41075161_R GTCTCGTGGGCTCGGTGTTGACAACAGCTCTGGCA 60.1 0.79
mm1__chr3 175.98 Mbp_chr3_175978501_F TCGTCGGCAGCGTCGAGTCCTTCTGACCCATGCC 60.1 0.70
mm1__chr3 175.98 Mbp_chr3_175978501_R GTCTCGTGGGCTCGGGCAAAGGCTCCTGTATGTGC 59.5 0.70
mm3__chr22 15.72 Mbp_chr22_15721541_F TCGTCGGCAGCGTCGTGCTCGTCATTGACAACGG 59.8 0.65
mm3__chr22 15.72 Mbp_chr22_15721541_R GTCTCGTGGGCTCGGATCATGTTCTGGTGCCTGGG 60.0 0.65
mm3__chr15 43.99 Mbp_chr15_43990106_F TCGTCGGCAGCGTCAATGAAGGGGAAGATGGCCC 59.7 0.65
mm3__chr15 43.99 Mbp_chr15_43990106_R GTCTCGTGGGCTCGGGCCAGCTCACCATGGATGAT 60.1 0.65
mm3__chr14 19.40 Mbp_chr14_19402830_F TCGTCGGCAGCGTCATCATGTTCTGGTGCCTGGG 60.0 0.65
mm3__chr14 19.40 Mbp_chr14_19402830_R GTCTCGTGGGCTCGGGTGCTCGTCATTGACAACGG 59.8 0.65
mm3__chr14 19.00 Mbp_chr14_18998922_F TCGTCGGCAGCGTCGTGCTCGTCATTGACAACGG 59.8 0.65
mm3__chr14 19.00 Mbp_chr14_18998922_R GTCTCGTGGGCTCGGATCATGTTCTGGTGCCTGGG 60.0 0.65
mm2__chr10 69.02 Mbp_chr10_69022808_F Primer3: not found at this Tm N.d. 0.65
mm2__chr10 69.02 Mbp_chr10_69022808_R Primer3: not found at this Tm N.d. 0.65
mm4__chr15 34.79 Mbp_chr15_34794724_F TCGTCGGCAGCGTCGGGAAGTTTACCTGGTGCCG 60.9 0.44
mm4__chr15 34.79 Mbp_chr15_34794724_R GTCTCGTGGGCTCGGATGTGTGACGACGAGGAGAC 59.4 0.44
mm4__chr17 44.12 Mbp_chr17_44123520_F Primer3: not found at this Tm N.d. 0.43
mm4__chr17 44.12 Mbp_chr17_44123520_R Primer3: not found at this Tm N.d. 0.43
mm4__chr19 57.63 Mbp_chr19_57633310_F TCGTCGGCAGCGTCTTCTGTCTGTCAGCCGCTTT 59.8 0.23
mm4__chr19 57.63 Mbp_chr19_57633310_R GTCTCGTGGGCTCGGGAACCCGAGCATCGCTATCC 60.6 0.23
mm4__chr3 50.15 Mbp_chr3_50154818_F TCGTCGGCAGCGTCGCGATGTGTGCGGTGTATTC 59.9 0.20
mm4__chr3 50.15 Mbp_chr3_50154818_R GTCTCGTGGGCTCGGGGGCTCAGGGTTCCCAGG 61.7 0.20
mm4__chr19 35.13 Mbp_chr19_35126560_F Primer3: not found at this Tm N.d. 0.16
mm4__chr19 35.13 Mbp_chr19_35126560_R Primer3: not found at this Tm N.d. 0.16
mm4__chr17 81.65 Mbp_chr17_81647955_F TCGTCGGCAGCGTCGAACTGACGATTCCATGCGC 59.9 0.16
mm4__chr17 81.65 Mbp_chr17_81647955_R GTCTCGTGGGCTCGGCACTGGTCGTTGACAGAGGC 60.9 0.16
mm4__chr10 88.95 Mbp_chr10_88948846_F Primer3: not found at this Tm N.d. 0.13
mm4__chr10 88.95 Mbp_chr10_88948846_R Primer3: not found at this Tm N.d. 0.13
mm4__chr7 152.68 Mbp_chr7_152675888_F TCGTCGGCAGCGTCAGGTGAGAGGTGGCAGAGG 60.6 0.13
mm4__chr7 152.68 Mbp_chr7_152675888_R GTCTCGTGGGCTCGGGGATTGGGAGGCTTGGGG 59.7 0.13
mm3__chr9 6.83 Mbp_chr9_6834510_F TCGTCGGCAGCGTCCCGCTACCAGCTCACCATAG 59.9 0.10
mm3__chr9 6.83 Mbp_chr9_6834510_R GTCTCGTGGGCTCGGACACGATGAAGGGGAAGACG 59.7 0.10
mm4__chr5 31.19 Mbp_chr5_31194057_F TCGTCGGCAGCGTCGTTGGGAGGGATGCTCTCAG 59.8 0.10
mm4__chr5 31.19 Mbp_chr5_31194057_R GTCTCGTGGGCTCGGTACCAGGAGGCGGAGGAG 59.7 0.10
mm4__chr19 18.79 Mbp_chr19_18788699_F TCGTCGGCAGCGTCAGTCTGCATGCTCGTGGC 60.4 0.09
mm4__chr19 18.79 Mbp_chr19_18788699_R GTCTCGTGGGCTCGGCAGGGCTCCTTCCAGTGC 60.0 0.09
mm4__chr16 67.91 Mbp_chr16_67908714_F TCGTCGGCAGCGTCTGGCTGTGCTGACTTGTTCT 59.8 0.08
mm4__chr16 67.91 Mbp_chr16_67908714_R GTCTCGTGGGCTCGGCTCGGGAAGGACCTTGCTTG 60.6 0.08
mm4__chrX 40.16 Mbp_chrX_40158340_F Primer3: not found at this Tm N.d. 0.07
mm4__chrX 40.16 Mbp_chrX_40158340_R Primer3: not found at this Tm N.d. 0.07
mm4__chr3 184.26 Mbp_chr3_184261159_F Primer3: not found at this Tm N.d. 0.07
mm4__chr3 184.26 Mbp_chr3_184261159_R Primer3: not found at this Tm N.d. 0.07
mm4__chr21 6.55 Mbp_chr21_6553830_F TCGTCGGCAGCGTCGGAGGCACAGGAAGAAGCTT 59.9 0.06
mm4__chr21 6.55 Mbp_chr21_6553830_R GTCTCGTGGGCTCGGCAGTCTGCCTCTGAGCCAAA 59.9 0.06
mm4__chr21 43.16 Mbp_chr21_43162152_F TCGTCGGCAGCGTCGGAGGCACAGGAAGAAGCTT 59.9 0.06
mm4__chr21 43.16 Mbp_chr21_43162152_R GTCTCGTGGGCTCGGCAGTCTGCCTCTGAGCCAAA 59.9 0.06
mm4__chr12 114.69 Mbp_chr12_114692899_F TCGTCGGCAGCGTCAACAGGCTCAGAGAGAACGC 60.0 0.06
mm4__chr12 114.69 Mbp_chr12_114692899_R GTCTCGTGGGCTCGGGCGGTGGGAGAGGAGAACTC 61.9 0.06
mm4__chr1 64.97 Mbp_chr1_64966711_F Primer3: not found at this Tm N.d. 0.06
mm4__chr1 64.97 Mbp_chr1_64966711_R Primer3: not found at this Tm N.d. 0.06
mm2__chr5 77.78 Mbp_chr5_77784941_F TCGTCGGCAGCGTCATATCACCGCGCTCGTCATT 59.9 0.06
mm2__chr5 77.78 Mbp_chr5_77784941_R GTCTCGTGGGCTCGGAACGATGGAGGGGAAGACTG 59.0 0.06
mm4__chr10 42.55 Mbp_chr10_42552727_F TCGTCGGCAGCGTCTCTCCCACCTCCAGGACAAG 60.5 0.06
mm4__chr10 42.55 Mbp_chr10_42552727_R GTCTCGTGGGCTCGGTTCTCCGGTACAGCCTGGTA 59.9 0.06
mm4__chr4 1.77 Mbp_chr4_1772961_F TCGTCGGCAGCGTCGTGAGCGCTGGAGGTGTTAT 60.1 0.05
mm4__chr4 1.77 Mbp_chr4_1772961_R GTCTCGTGGGCTCGGATTTCCAGTTGGCCCCATCC 60.3 0.05
mm4__chr17 19.65 Mbp_chr17_19648198_F TCGTCGGCAGCGTCGTGCGAGGATGATGCTTGTG 59.6 0.04
mm4__chr17 19.65 Mbp_chr17_19648198_R GTCTCGTGGGCTCGGTTCCCTGGAGAGCTAGTCCC 60.0 0.04
mm4__chr5 98.34 Mbp_chr5_98338798_F TCGTCGGCAGCGTCGCTCCTCATCAACAAGGGCT 60.0 0.04
mm4__chr5 98.34 Mbp_chr5_98338798_R GTCTCGTGGGCTCGGTTTTGACCCATGCCCACCAT 60.1 0.04
mm4__chr1 229.43 Mbp_chr1_229433031_F TCGTCGGCAGCGTCCGATGGACGGGAACACGG 60.5 0.03
mm4__chr1 229.43 Mbp_chr1_229433031_R GTCTCGTGGGCTCGGGTCTCCCTGTCCTTGCAGAA 59.3 0.03
mm4__chr19 50.40 Mbp_chr19_50402330_F TCGTCGGCAGCGTCTGGAACAGGGCATCCGTG 59.6 0.03
mm4__chr19 50.40 Mbp_chr19_50402330_R GTCTCGTGGGCTCGGCCGGATCTCAAAGTCGACGT 59.8 0.03
mm4__chr7 3.84 Mbp_chr7_3842288_F TCGTCGGCAGCGTCATCCTTCCTCTCCCCAGACC 60.0 0.02
mm4__chr7 3.84 Mbp_chr7_3842288_R GTCTCGTGGGCTCGGGGCACCCTTGGGTATTCTCC 60.1 0.02
mm4__chr13 99.90 Mbp_chr13_99896062_F Primer3: not found at this Tm N.d. 0.00
mm4__chr13 99.90 Mbp_chr13_99896062_R Primer3: not found at this Tm N.d. 0.00
mm4__chr11 134.41 Mbp_chr11_134412445_F TCGTCGGCAGCGTCCGGGATTGGTATCACCTCCC 59.6 0.00
mm4__chr11 134.41 Mbp_chr11_134412445_R GTCTCGTGGGCTCGGTCGAGAGCCGAGATCTGGG 60.5 0.00
mm2__chr5 131.66 Mbp_chr5_131660037_F TCGTCGGCAGCGTCATGCCCATCATCACACCCTG 60.1 0.00
mm2__chr5 131.66 Mbp_chr5_131660037_R GTCTCGTGGGCTCGGTGTCGTCGACAATGGCTCTG 60.3 0.00
mm3__chr11 1.80 Mbp_chr11_1804017_F TCGTCGGCAGCGTCCGCCACCCACATAGAAGTCC 60.4 0.00
mm3__chr11 1.80 Mbp_chr11_1804017_R GTCTCGTGGGCTCGGAACACCTCTGGCATGTGCAA 60.4 0.00
mm4__chr15 88.71 Mbp_chr15_88705601_F TCGTCGGCAGCGTCCTCGGGCTCTCATTTACCCC 59.8 0.00
mm4__chr15 88.71 Mbp_chr15_88705601_R GTCTCGTGGGCTCGGCTGCCCAGTCCGGTCTGC 62.7 0.00
mm4__chr2 109.62 Mbp_chr2_109615229_F TCGTCGGCAGCGTCCTCAGACAGCGCATCGGT 59.8 0.00
mm4__chr2 109.62 Mbp_chr2_109615229_R GTCTCGTGGGCTCGGCTCCAGGCTGTCCCACTTG 60.0 0.00
mm4__chr3 12.81 Mbp_chr3_12810290_F TCGTCGGCAGCGTCCATCCGCACCCTGATCCAAT 60.1 0.00
mm4__chr3 12.81 Mbp_chr3_12810290_R GTCTCGTGGGCTCGGCGCCCCTTTACCGAGTCATT 60.1 0.00
mm4__chr13 44.93 Mbp_chr13_44930531_F Primer3: not found at this Tm N.d. 0.00
mm4__chr13 44.93 Mbp_chr13_44930531_R Primer3: not found at this Tm N.d. 0.00
mm2__chr3 180.82 Mbp_chr3_180820598_F TCGTCGGCAGCGTCAATGATATCGCCACGCTCGT 59.9 0.00
mm2__chr3 180.82 Mbp_chr3_180820598_R GTCTCGTGGGCTCGGTTCTTCTGACCCATGCCCAC 59.9 0.00

Off-target amplicon sequences

Primers underlined, off-targets in bold.

ontarget_mm0__chr7 5.53 Mbp_chr7_5529579 CACGATGGAGGGGAAGACGGCCCGGGGGGCATCGTCGCCCGCGAAGCCGGCCTTGCACATGCCGGAGCCGTTGTCGACGA
CGAGCGCGGCGATATCATCATCCATGGTGAGCTGCGAGAATAGC
mm1__chr2 131.63 Mbp_chr2_131626193 GTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCGGGCGACGATGCCCCCCAGGCTGTCTTCCCTTC
TATCGTGGGGCGCCCCAGGCACCAGGGCATGATGGAGGGCATGCATCAGAAGGAGT
mm1__chr2 131.26 Mbp_chr2_131263610 TGGATGATGATACCGCCGTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCGGGCGACGATGCCCCC
CGGGCTGTCTTCCCTTCCATC
mm1__chr2 130.66 Mbp_chr2_130656804 GTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCGGGCGACGATGCCCCCCGGGCTGTCTTCCCTTC
CATC
mm1__chr2 130.46 Mbp_chr2_130463865 GATGGAAGGGAAGACAGCCCGGGGGGCATCGTCGCCCGCAAAGCCGGCCTTGCACATGCCAGAGCCGTTGTCAATGACGA
GCAC
mm1__chr2 130.08 Mbp_chr2_130075293 GATGGAAGGGAAGACAGCCCGGGGGGCATCGTCGCCCGCAAAGCCGGCCTTGCACATGCCAGAGCCGTTGTCAATGACGA
GCAC
mm2__chr22 41.08 Mbp_chr22_41075161 GGAGTCCTTCTGACCCATGCCCACCATCATGACCTGGTGCCTGGTGCACCCCACGATGGAAGGGAAGATGGCCCAGGGGA
CATCGTCACCCGCGAAGCCGGCCTTGCACATGCCAGAGCTGTTGTCAACA
mm1__chr3 175.98 Mbp_chr3_175978501 GAGTCCTTCTGACCCATGCCCACCATCACACCTTGGTGCCTGGGGCGCCCCACGATGGATGGGAAGACGGCCCCGGAGGC
ATCGTCGCCTGCGAAGCTGGTCTTGCACATACAGGAGCCTTTGC
mm3__chr22 15.72 Mbp_chr22_15721541 GTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCAGGTGACGATGCCCCCCGGGCTGTCTTCCCTTC
CATCGTGGGGTGCCCCAGGCACCAGAACATGAT
mm3__chr15 43.99 Mbp_chr15_43990106 AATGAAGGGGAAGATGGCCCGGGGTGGGCATCGTCACCTGCAAAGCTGGCCTTGCACATGCCAGAGCCATTGTCGACGAC
AAGCATGGTGATATCATCATCCATGGTGAGCTGGC
mm3__chr14 19.40 Mbp_chr14_19402830 ATCATGTTCTGGTGCCTGGGGCACCCCACGATGGAAGGGAAGACAGCCCGGGGGGCATCGTCACCTGCAAAGCCGGCCTT
GCACATGCCAGAGCCGTTGTCAATGACGAGCAC
mm3__chr14 19.00 Mbp_chr14_18998922 GTGCTCGTCATTGACAACGGCTCTGGCATGTGCAAGGCCGGCTTTGCAGGTGACGATGCCCCCCGGGCTGTCTTCCCTTC
CATCGTGGGGTGCCCCAGGCACCAGAACATGAT
mm4__chr15 34.79 Mbp_chr15_34794724 GGGAAGTTTACCTGGTGCCGCGGGCGGCCCACGATGGACGGGAAGACAGCGCGGGGCGCGTCATCGCCCGCAAAGCCGGC
CTTCACCAGCCCAGAGCCGTTGTCGCACACCAGGGCGGTGGTCTCCTCGTCGTCACACAT
mm4__chr19 57.63 Mbp_chr19_57633310 TTCTGTCTGTCAGCCGCTTTGGTACGCTGCATCGGGATCGAAGTGACGGACCGTGAAGGCGCGAGAGTCAGGTCTGAGGG
TCGGGGGCAGAGCCGCCCGCGAGGCCGGCCTGGGGATAGCGATGCTCGGGTTC
mm4__chr3 50.15 Mbp_chr3_50154818 GCGATGTGTGCGGTGTATTCACAGAGAGACAAAGCCANNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNN
NNNGTGCAGACTAGTCTCCCCCGGCGCGGGGTGCCCACCGCCTGGGAACCCTGAGCCC
mm4__chr17 81.65 Mbp_chr17_81647955 GAACTGACGATTCCATGCGCCTTGCAGGTACTTTAACTGCAGCTCCCCCGGGGTGCAGGCCTGCAGCCTTCCCGCCTCCT
GCTGCATCGACCCCCGCGAAGATGGAGCCTCTGTCAACGACCAGTG
mm4__chr7 152.68 Mbp_chr7_152675888 AGGTGAGAGGTGGCAGAGGGTGCCAGCATCGCGGGCGTCTAGGCCGAGAGGCCGGTCCCAGCCCGGCCAGGCCGCGCCGC
CCCAAGCCTCCCAATCC
mm3__chr9 6.83 Mbp_chr9_6834510 CCGCTACCAGCTCACCATAGATGATGATATTGCTACGCCCGTCGTCGACAACGGCTCCGGCATGTGGAAGGCTGGCTTCG
CAGGTGACGATGACCCCCGGCCGTCTTCCCCTTCATCGTGT
mm4__chr5 31.19 Mbp_chr5_31194057 GTTGGGAGGGATGCTCTCAGCCTNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNNGCGCTGCTGCCG
CTGCTCAGGACCGACGCCGCTTCCCGGGGAGCCTGGGCACCGCTCCTCCGCCTCCTGGTA
mm4__chr19 18.79 Mbp_chr19_18788699 AGTCTGCATGCTCGTGGCACTCGCTGGGCGAGCCGTCGGGGCAGAAGCGCTGTGCGCGCCGCTGGCAGCCGGACGCCTGG
TCGCCCACGAAGCCGGGCTGGCACGGGCCGCACTGGAAGGAGCCCTG
mm4__chr16 67.91 Mbp_chr16_67908714 TGGCTGTGCTGACTTGTTCTCTCTTTGTGTGTAGGTGTAGACGGGGCACTGCCTTCAGAGCAGGTCCTGCCAGCCTCGCT
GGAGAGGATGCCCTCGTGTCCGTGATGGGCTGTGGGACAAGCAAGGTCCTTCCCGAG
mm4__chr21 6.55 Mbp_chr21_6553830 GGAGGCACAGGAAGAAGCTTACCTCACCCTTCAGACATCCTGGGGGGTACCCTTGTTTCCTATATCATTTCAACCTTGGG
GTCCCAGCATTGCTGGCGACGAGGCCGGCTGCAGGTTTTGGCTCAGAGGCAGACTG
mm4__chr21 43.16 Mbp_chr21_43162152 GGAGGCACAGGAAGAAGCTTACCTCACCCTTCAGACATCCTGGGGGGTACCCTTGTTTCCTATATCATTTCAACCTTGGG
GTCCCAGCATTGCTGGCGACGAGGCCGGCTGCAGGTTTTGGCTCAGAGGCAGACTG
mm4__chr12 114.69 Mbp_chr12_114692899 AACAGGCTCAGAGAGAACGCAGCCGAGCGGGGGAATCTTCGTGCGCGAAGCCGGGAGGGTTGCGGGGAGGGAGAGTTCTC
CTCTCCCACCGC
mm2__chr5 77.78 Mbp_chr5_77784941 ATATCACCGCGCTCGTCATTGACAACGGCTCCGGCATGTGCAAGGCCAGCTTCACGGGCGACAATGCCGCCCGGGCAGTC
TTCCCCTCCATCGTT
mm4__chr10 42.55 Mbp_chr10_42552727 TCTCCCACCTCCAGGACAAGCGGCACCTCCAGTCCACCCTCGCTCCAGCCCTCCCGTACTCTACCCCCGGCATCCTCTGC
CCCGAAGCAGGGAGATGCCGGGAGCCCTCCCTACCAGGCTGTACCGGAGAA
mm4__chr4 1.77 Mbp_chr4_1772961 GTGAGCGCTGGAGGTGTTATCTCTGACAGGAGCAGAATGGACTCGTCGCGCCCGAAGCAGGTGTCGGCGCCTGGGATGGG
GCCAACTGGAAAT
mm4__chr17 19.65 Mbp_chr17_19648198 GTGCGAGGATGATGCTTGTGCAAGAAGGAAATGACCGCTGTCCCGGCTCCGCGGGAGACGAACCCCGGCTTCCCGCCCTC
AGGGACTAGCTCTCCAGGGAA
mm4__chr5 98.34 Mbp_chr5_98338798 GCTCCTCATCAACAAGGGCTCTGGCATGTGCAAGGCCGGCTTCACAGGTGACAATGCCCCCGGGCCATCTTCCCCTCTGT
CATGGGTGCCAGGGAGTGATGGTGGGCATGGGTCAAAA
mm4__chr1 229.43 Mbp_chr1_229433031 CGATGGACGGGAACACGGCCCTAGGGGCGTCATCCCCGGCGAAGCCGGCTTTCACCAGGCCGGAGCCATTGTCGCACACG
AGGGCGGTGGTCTCGTCTTCGTCGCACATTGTGTCTAGTTTCTGCAAGGACAGGGAGAC
mm4__chr19 50.40 Mbp_chr19_50402330 TGGAACAGGGCATCCGTGTGGCAGGCCTGGGCACGCCCAGCTTCGCGCCCTACGAGGCCAACGTCGACTTTGAGATCCGG
mm4__chr7 3.84 Mbp_chr7_3842288 ATCCTTCCTCTCCCCAGACCTTAAGCTCTGCTGTGTTAGAGTCCTGCATCCCGGGAGATGATGCCTCCACTGGAGAATAC
CCAAGGGTGCC
mm4__chr11 134.41 Mbp_chr11_134412445 CGGGATTGGTATCACCTCCCCACCGCCCCCCACCCTCCCCCGGCACCGTCCCCCGCGGAGGCGGCTCCCGCTTCAGGCGG
CGGAGCAGGGGTGGCGGGCAGGGGCCGAGCGCACCCCCAGATCTCGGCTCTCGA
mm2__chr5 131.66 Mbp_chr5_131660037 ATGCCCATCATCACACCCTGGTGCCTCGGGTGCCCCATTATGGAGGGGAAGACGGCCCGGGGGCATCGTTGCCCGTGAAG
CAGACCTTCCACATGCCAGAGCCATTGTCGACGACA
mm3__chr11 1.80 Mbp_chr11_1804017 CGCCACCCACATAGAAGTCCTTCTGACCCATGCCCACCGTCATGCCCTGGTGTCTGGGGCACCCCACGATGGAGGGGAAG
ACGGCCCAGGGGGCATCGTCGTCGGTGAAGCTGGCCTTGCACATGCCAGAGGTGTT
mm4__chr15 88.71 Mbp_chr15_88705601 CTCGGGCTCTCATTTACCCCGGCCGCTGGCAGCGGGACCCAGGCGCCTTGCCGCCTCGCTGCGGCCGCTTGGTCCCCACG
CCGCCCCCTCCGCAGGCGACGAGGCCTGCAGACCGGACTGGGCAG
mm4__chr2 109.62 Mbp_chr2_109615229 CTCAGACAGCGCATCGGTGGCCTCGTCGTCCGCGGAGGAGGAGAGCAGCGGCGGAGGCTCCGTGACGCTGGACCCCCTGG
AGCACGCGTGGATGCTCTCTGCCTCCGATGGCAAGTGGGACAGCCTGGAG
mm4__chr3 12.81 Mbp_chr3_12810290 CATCCGCACCCTGATCCAATGTTTGGGCAGCGTCGGCCGCCAGGCCGGCCACCGCCTCGGTAAGGGGGCAGGGGGCGGGG
CCTGGGCTGGCATGGTGGGCGGAGCTTAGGGTGAGCCAATGACTCGGTAAAGGGGCG
mm2__chr3 180.82 Mbp_chr3_180820598 AATGATATCGCCACGCTCGTCATGGACAATGGCTCTCCCATGTGCAAGGCCAGCTTAGCAGGCGACGATGCCCCTCCATC
GTGAGGCACCCATGGCACCAGGGCATGATCGTGGGCATGGGTCAGAAGAA

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.