CRISPR Library Design Pipeline: Difference between revisions
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#Create a manually curated gene list for each library. | #Create a manually curated gene list for each library. | ||
#Pick 3-4 gRNAs per gene with the highest score from this [[File:gRNA_spacer_file.txt]] | #Pick 3-4 gRNAs per gene with the highest score from this [[File:gRNA_spacer_file.txt]] | ||
#Add scaffold sequences and desired barcodes. The barcode list can be found in [[ | #Add scaffold sequences and desired barcodes. The barcode list can be found in [[File:gRNA_bc25mers.txt]]. The scaffold sequences can be found on this page: [[gRNA Scaffold Sequences]]. | ||
The process for designing the [[05/30/2016 Library Order]] includes: | |||
#Create a manually curated gene list for each library | |||
#Sort gRNAs from this list: [[File:Exon_Filtered_gRNA_Spacer_File.txt]] by exon first, and then score. Pick 3 gRNAs per gene for a single library. | |||
#For a double library, take combinations of 2 gRNAs per gene using the highest ranked genes most often. For example, if we have 4 gRNAs ranked gRNA_1, gRNA_2, gRNA_3, gRNA_4, the dual combinations would be (gRNA_1, gRNA_2), (gRNA_1, gRNA_3), (gRNA_1, gRNA_4). | |||
#Add scaffold sequences and desired barcodes. The barcode list can be found in [[File:trimmed_bc25mers.txt]]. These barcodes are 18 bp instead of 25 bp in order to accommodate the dual libraries. They've been filtered so that all bc25mers are at least an edit distance of 9 apart. | |||
The code for making libraries and checking to make sure gRNA spacers map to the expected gene/exon is attached here: [[File:CRISPR_Library_Construction.zip]] |
Latest revision as of 23:27, 21 May 2016
The process for designing the 05/05/2015 Library Order and 02/17/2016 Library Order included:
- Create a manually curated gene list for each library.
- Pick 3-4 gRNAs per gene with the highest score from this File:GRNA spacer file.txt
- Add scaffold sequences and desired barcodes. The barcode list can be found in File:GRNA bc25mers.txt. The scaffold sequences can be found on this page: gRNA Scaffold Sequences.
The process for designing the 05/30/2016 Library Order includes:
- Create a manually curated gene list for each library
- Sort gRNAs from this list: File:Exon Filtered gRNA Spacer File.txt by exon first, and then score. Pick 3 gRNAs per gene for a single library.
- For a double library, take combinations of 2 gRNAs per gene using the highest ranked genes most often. For example, if we have 4 gRNAs ranked gRNA_1, gRNA_2, gRNA_3, gRNA_4, the dual combinations would be (gRNA_1, gRNA_2), (gRNA_1, gRNA_3), (gRNA_1, gRNA_4).
- Add scaffold sequences and desired barcodes. The barcode list can be found in File:Trimmed bc25mers.txt. These barcodes are 18 bp instead of 25 bp in order to accommodate the dual libraries. They've been filtered so that all bc25mers are at least an edit distance of 9 apart.
The code for making libraries and checking to make sure gRNA spacers map to the expected gene/exon is attached here: File:CRISPR Library Construction.zip