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Amplicon sequencing of STING gene to validate CRISPR-Cas9 cut sites in PC9 cell line

ABSTRACT: Cells carrying lentiCRISPRv2 e.v. or lentiCRISPRv2 sgSTING were selected with puromycin before DNA was extracted to assess the genomic alterations at the target locus.

INSTRUMENT(S): Illumina NovaSeq 6000

ORGANISM(S): Homo sapiens

SUBMITTER: Johannes Braegelmann

PROVIDER: E-MTAB-10851 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Amplicon sequencing of STING gene to validate CRISPR-Cas9 cut sites in PC9 cell line

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Project description:Elucidating the role of gut microbiota in physiological and pathological processes has recently emerged as a key research aim in life sciences. In this respect, metaproteomics (the study of the whole protein complement of a microbial community) can provide a unique contribution by revealing which functions are actually being expressed by specific microbial taxa. However, its wide application to gut microbiota research has been hindered by challenges in data analysis, especially related to the choice of the proper sequence databases for protein identification. Here we present a systematic investigation of variables concerning database construction and annotation, and evaluate their impact on human and mouse gut metaproteomic results. We found that both publicly available and experimental metagenomic databases lead to the identification of unique peptide assortments, suggesting parallel database searches as a mean to gain more complete information. Taxonomic and functional results were revealed to be strongly database-dependent, especially when dealing with mouse samples. As a striking example, in mouse the Firmicutes/Bacteroidetes ratio varied up to 10-fold depending on the database used. Finally, we provide recommendations regarding metagenomic sequence processing aimed at maximizing gut metaproteome characterization, and contribute to identify an optimized pipeline for metaproteomic data analysis.

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Combining transcriptome and proteome analysis to identify and characterize the RpoS regulon in Pseudomonas fluorescens

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