PRJDB4492 SAMD00045501 rnh201_CR_replicate1_mock-treatment_Background OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045502 rnh201_CR_replicate1_mock-treatment_Nick OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045503 rnh201_CR_replicate1_mock-treatment_Nick_PCR-amplfied OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045504 rnh201_CR_replicate1_RNaseH-digested_Background OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045505 rnh201_CR_replicate1_RNaseH-digested_rNMP OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045506 rnh201_CR_replicate1_RNaseH-digested_rNMP_PCR-amplfied OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045507 rnh201_CR_replicate2_RNaseH-digested_Background OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045508 rnh201_CR_replicate2_RNaseH-digested_rNMP OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq PRJDB4492 SAMD00045509 rnh201_CR_replicate2_RNaseH-digested_rNMP_PCR-amplfied OTHER GENOMIC other "RisCQ-seq was performed by non-separated library preparation from genomic DNA. DNA was isolated from yeast spheloplasts by lysis buffer (50 mM Tris-Cl pH7.5, 0.2 M NaCl, 0.1 M EDTA and 5% SDS) and phenol-chloroform- isoamylalcohol extraction prior to 0.5 mg/ml RNaseA treatment. Fifty _g of total DNA was sheared in microTUBE (Covaris) by Covaris M220 (peak power 140, duty factor 10, cycle/burst 200 and duration 1 minute). Agarose-gel purified DNA (400-500 bp range) was end-repaired by KAPA library preparation kit for Illumina (Kapa Biosystems) supplement with 2 mM ATP and 2.5 mM dNTP. One _g of the size-selected DNA and 0.1 _g of spike-in standard cocktail were used for a primary library. Pre-primary library was prepared by KAPA library preparation kit and Illumina TruSeqHT-compatible adaptors (IDT). Free 3'-OH ends of the pre-primary library were masked by terminal deoxynucleotidyl transferase (Takara) with 0.25 mM ddNTP (Roche) for 1 hour at 37‹C. The fully ligated library was purified by size-selection with agarose-gel electrophoresis (primary library). For RNaseH-digested samples, 100 ng of the primary library was digested using 1xThermoPol reaction buffer, 3.75 units of RNase H and RNase HII (New England Biolabs) overnight (16 hours) at 37‹C. Thirty ng of the primary library with or without RNaseH digestion was heat denatured at 100‹C for 3 minutes and chilled at 0‹C for 5 minutes. The denatured library was incubated with 10 _M of 701ss adaptor and 750 units of T4 DNA ligase (Kapa Biosystems) overnight (12 hours) at 20‹C for secondary-adaptor ligation. After three times of library purification with AMPureXP beads (Beckman Coulter), complementary strand of the libraries were synthesized in 25 _l reaction mixture with 16 units of BstPolymerase LF, 1xThermoPol reaction buffer (New England Biolabs), 0.25 mM dNTP and 4 _M of TruHTAmp7Rv primer at 65‹C for 10 minutes. The size-selected libraries were directly used for paired-end sequencing by MiSeq sequencer (Illumina) to identify location of fragments and 3'-end of nicks generated with or without RNaseH treatment. For PCR-amplified sample, 5 ng of the double-stranded libraries were amplified using 5 _M of primers TruHTAmp5Fw and TruHTAmp701allSRv, using KAPA real time library amplification kit (Kapa Biosystems). Libraries were then purified with AMPureXP beads (target range over 300 bp) and sequenced as PCR-amplified samples." 150 0 Application Read Forward 1 1 Application Read Reverse 76 Illumina MiSeq