CRISPR screen and single-cell sequencing project: Difference between revisions
Jump to navigation
Jump to search
>YanWu No edit summary |
>YanWu No edit summary |
||
Line 1: | Line 1: | ||
The goal of this project is to screen H9-GFP tagged hESCs with 4 curated libraries and then use single-cell sequencing to determine which gRNA was incorporated into which cell and gene expression in each cell. The four libraries and some annotation info is listed [[File:Crispr_screen_single_cell_sequencing_library.xlsx]]. | The goal of this project is to screen H9-GFP tagged hESCs with 4 curated libraries and then use single-cell sequencing to determine which gRNA was incorporated into which cell and gene expression in each cell. The four libraries and some annotation info is listed in this file ([[File:Crispr_screen_single_cell_sequencing_library.xlsx]]). | ||
Library Design: | Library Design: | ||
# | #From the curated gene lists, take all spacers for each gene from the gRNA list. | ||
#Check each spacer for multi-mapping and that each spacer maps to the correct location. | |||
#Keep only the top 3 spacers for each gene. | |||
#Include 3 negative controls in each library, barcode using bc_25mer barcodes. | |||
#Order gChip. See [[05/05/2015 Library Order]]. | |||
#Use [[CRISPR Library Prep Protocol]] to prepare library. |
Revision as of 00:23, 6 May 2015
The goal of this project is to screen H9-GFP tagged hESCs with 4 curated libraries and then use single-cell sequencing to determine which gRNA was incorporated into which cell and gene expression in each cell. The four libraries and some annotation info is listed in this file (File:Crispr screen single cell sequencing library.xlsx).
Library Design:
- From the curated gene lists, take all spacers for each gene from the gRNA list.
- Check each spacer for multi-mapping and that each spacer maps to the correct location.
- Keep only the top 3 spacers for each gene.
- Include 3 negative controls in each library, barcode using bc_25mer barcodes.
- Order gChip. See 05/05/2015 Library Order.
- Use CRISPR Library Prep Protocol to prepare library.