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Series GSE179614 Query DataSets for GSE179614
Status Public on Feb 24, 2022
Title Rapid cell-free characterization of multi-subunit CRISPR effectors and transposons
Organisms Escherichia coli; synthetic construct
Experiment type Other
Summary We report the PAMs of diverse type I-E CRISPR- Cas systems and the type I-C and the type I-F1 CRISPR-Cas systems from Xanthomonas albilineans. Furthermore, we report PAMs of two type I-B CRISPR transposons (CASTs) and the Vibrio cholerae type I-F CAST. For identification of the PAMs, we used a cell-free TXTL-based PAM screen we named PAM-DETECT. By adding a 5N randomized PAM library and plasmids encoding for Cascade genes and gRNAs, recognized PAMs were bound by Cascade and protected from cleavage by a restriction enzyme that has it's recognition site within the target region. By amplifying the non-cleaved target plasmid, we used next-generation sequencing to analyze the enrichment of functional PAMs of the studied CRISPR-Cas systems. We additionally assessed the insertion sites of crRNA-dependent and crRNA-independent transposition of the Rippkaea orientalis type I-B CAST in TXTL and E. coli.
 
Overall design PAMs for 12 different type I-E CRISPR systems, one type I-C, one type I-F1 system, two type I-B CASTs and one type I-F1 CAST were determined. Insertion points for crRNA-dependent and crRNA-independent transposition were assessed for one type I-B CAST.
 
Contributor(s) Wimmer F, Mougiakos I, Englert F, Beisel CL
Citation(s) 35216669
Submission date Jul 06, 2021
Last update date May 29, 2022
Contact name Chase Lawrence Beisel
Organization name Helmholtz Centre for Infection Research (HZI)
Department Helmholtz-Institute for RNA-based Infection Research (HIRI)
Lab RSYN
Street address Josef-Schneider Straße 2
City Würzburg
ZIP/Postal code 97080
Country Germany
 
Platforms (3)
GPL16085 Illumina MiSeq (Escherichia coli)
GPL17769 Illumina MiSeq (synthetic construct)
GPL26526 Illumina NovaSeq 6000 (synthetic construct)
Samples (89)
GSM5425007 Ec_3nM_16h_digested_rep1
GSM5425008 Ec_3nM_16h_digested_rep2
GSM5425009 Ec_3nM_16h_undigested_rep1
Relations
BioProject PRJNA744274
SRA SRP327230

Download family Format
SOFT formatted family file(s) SOFTHelp
MINiML formatted family file(s) MINiMLHelp
Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE179614_RAW.tar 820.0 Kb (http)(custom) TAR (of TXT)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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