Cas9 targeted enrichment of mobile elements using nanopore sequencing in human genomes

Nano-Pal pipeline for Nanopore reads from Flongle/MinION

Run for L1Hs

bash Nano-Pal.LINE.sh

Run for AluYb

bash Nano-Pal.AluYb.sh

Run for AluYa

bash Nano-Pal.AluYa.sh

Run for SVA_F

bash Nano-Pal.SVA_F.sh

Run for SVA_E

bash Nano-Pal.SVA_E.sh

Requires:

 samtools/1.3.1  https://github.com/samtools/samtools
 ncbi-blast++/2.10.0  ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST/
 PALMER https://github.com/mills-lab/PALMER
 minimap2 https://github.com/lh3/minimap2

Guide RNA design pipeline

We have included three bash files for guide RNA design: bash.alu.sh, bash.line1.sh, and bash.sva.sh in the RNA.design.pipelines folder.

We also included the consensus sequences for different categories of mobile elements used for the design in the lib folder.

Requires:

 jellyfish/2.2.8

Guide RNA cleavage-site analysis

Run MEI_ONTreads_alignment.py to perform alignments.
Accepted mobile element names: L1HS, AluYa5, AluYb8, SVA_E, SVA_F

MEI_ONTreads_alignment.py {mobileElementName} {mobileElementSeq.fasta} {long_reads_input_file.fasta} {outfile.txt}

Requires:

 import sys 
 import Bio

L1Hs methylation analysis

We have included two scripts for L1Hs methylation analysis: non_ref_pipeline.sh and reference_piepline.sh in the Methylation.pipelines folder.

Requires:

 nanopolish https://github.com/jts/nanopolish
 methylartist https://github.com/adamewing/methylartist

Citation

• Torrin L. McDonald*, Weichen Zhou*, Christopher Castro, Camille Mumm, Jessica A. Switzenberg, Ryan E. Mills, Alan P. Boyle, Cas9 targeted enrichment of mobile elements using nanopore sequencing, Nature Communications, 2021, https://doi.org/10.1038/s41467-021-23918-y

For PALMER:

• Weichen Zhou, Sarah B Emery, Diane A Flasch, Yifan Wang, Kenneth Y Kwan, Jeffrey M Kidd, John V Moran, Ryan E Mills, Identification and characterization of occult human-specific LINE-1 insertions using long-read sequencing technology, Nucleic Acids Research, 2019, https://doi.org/10.1093/nar/gkz1173

Contact:

For Nano-Pal and others: [email protected] or https://github.com/WeichenZhou

For cleavage-site anlaysis: [email protected] or https://github.com/castrocp

For methylation analysis: [email protected] or https://github.com/crmumm