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Sample GSM8016785

Query DataSets for GSM8016785

Status

Public on Jan 19, 2024

Title

WT2_M2G [Tn-seq]

Sample type

SRA

Source name

Colonies

Organism

Caulobacter vibrioides

Characteristics

cell type: Colonies
genotype: WT
treatment: Colonies grown on M2G

Growth protocol

A mini-Tn5 was introduced into C. crescentus WT by bi-parental mating on PYE agar for 4h at 30 ° C. Cells were collected, resuspended in 10 ml M2G or M2G-K and washed twice with the same volume. Dilutions (10-6) were plated on M2G or M2G-K plates supplemented with aztreonam (5mg l-1) and kanamycin (5mg l-1) and incubated at 30 ° C for 5 days. Then, at least 3 x10^5 colonies were collected in M2G or M2G-K supplemented with 10% glycerol and stored at -80 °C.

Extracted molecule

genomic DNA

Extraction protocol

Genomic DNA was then extracted following the Nucleospin Tissue Kit protocol (Macherey-Nagel) and resuspended in 50 µl elution buffer (5 mM Tris-HCl [pH 8.5]).
Tn-Seq libraries were prepared at the Fasteris company (Switzerland).

Library strategy

OTHER

Library source

genomic

Library selection

other

Instrument model

Illumina NextSeq 500

Data processing

*library strategy: Tn-seq
Around 3 x 10^7 paired-end sequence reads (2 x 75) were first filtered with the 5'-GGTTGAGATGTGTATAAGAGACAG sequence (corresponding to the mini-Tn5) and mapped on the genome of C. crescentus NA1000 (NC_011916.1) and converted to SAM using BWA and SAM tools from the sourceforge server (https://sourceforge.net/)
the number of reads overlapping each genomic position was computed using custom Python scripts.
The total number of reads for internal 80% of each ORF in each condition were then computed using custom Python scripts.
For comparing strains, the total number of reads was normalized by the ratio of the number of reads between the two strains.
Ward’s clustering analysis was performed to define the fitness categories as previously reported.
Assembly: NC_011916.1
Supplementary files format and content: The xls file reports Tn-Seq data. The different columns are organized as follows: column 1: Gene_ID (CCNA gene identification number); column 2: Start (coordinates for the start of the gene); column 3: End (coordinates for the end of the gene); column 4: Strand (Strand on which the gene is encoded); column 5: Name (Predicted function of the corresponding gene);[columns 6-13: WT_M2G; columns 14-21: WT_M2G-K]; columns 6&14: #Reads (number of normalized counts between the coordinates of the gene); columns 7&15: #Reads/bp (number of normalized counts divided by the length of the gene); columns 8&16: #Insertions (number of independent Tn insertions into the corresponding gene); columns 9&17: #Insertions/bp (number of independent Tn insertions divided by the length of the corresponding gene); columns 10&18: ES,#Reads (number of normalized counts between 80% of the coordinates of the gene); columns 11&19: ES,#Reads /bp (number of normalized counts divided by the 80% of length of the gene); columns 12&20: ES,#Insertions (number of independent Tn insertions into 80% of the corresponding gene); columns 13&21: ES,#Insertions/bp (number of independent Tn insertions divided by the 80% length of the corresponding gene)

Submission date

Jan 14, 2024

Last update date

Jan 19, 2024

Contact name

Regis Hallez

E-mail(s)

[email protected]

Organization name

UNamur

Street address

61 Rue de Bruxelles