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Sample GSM2242250

Query DataSets for GSM2242250

Status

Public on Oct 25, 2017

Title

Lib2_AAV

Sample type

SRA

Source name

AAV

Organism

Adeno-associated virus

Characteristics

tissue: AAV

Extracted molecule

other

Extraction protocol

Phenol-Chloroform
AAV barcodes were amplified with KAPA HIFI Hotstart using Primer #1 and indexing primers

Library strategy

OTHER

Library source

genomic

Library selection

other

Instrument model

Illumina HiSeq 2500

Description

DNA Barcodes in AAV

Data processing

Library strategy: Parallel Reporter Assay
A custom pipeline was created to assign barcodes to each tested sequences. Then sequence activity was calculated as the ratio of barcode abundance in the RNA sample compare to its abundance in the AAV pool.
Barcode sequences were extracted from 50bp reads by taking only reads starting with the expected backbone sequence: “TCCTGCTGGAGTTCGTGACCTGCATGCGCCGAA”. From these reads the sequence at position 34-48 was extracted. The frequency of each barcode sequence was calculated to get counts for each sample. Counts of barcodes were normalized to library size. Enrichment of barcodes in the RNA sample was calculated over their representation in the AAV input. Barcodes not sufficiently covered in the AAV sequencing were discarded (depending on sequencing depth 2-16 reads). The median activity of all barcodes per CRE was calculated. Only CREs that were covered in at least 2 out of 3 biological replicates with at least three barcodes were used for downstream analysis.
Genome_build: mm9

Submission date

Jul 19, 2016

Last update date

May 15, 2019

Contact name

Dirk Schuebeler

Organization name

Friedrich Miescher Institute for Biomedical Research

Street address

Maulbeerstrasse 66

City

Basel