Project description:Here we report the massively parallel cDNA sequencing (RNA-seq) analysis performed using high throughput sequencing of wild type (DB110) and toxR (TW30) mutant strains of the deep-sea bacterium Photobacterium profundum. ToxR is a transmembrane DNA-binding protein first discovered in Vibrio cholerae and able to regulate numerous genes involved in virulence. In P. profundum the abundance and activity of the same protein is influenced by hydrostatic pressure and is able to regulate genes in a pressure-dependent manner. To better characterize the ToxR regulon, we have compared the genes differentially expressed in response to pressure changes with those whose expression is altered between wild type and toxR mutant strains.
Project description:Here we report the massively parallel cDNA sequencing (RNA-seq) analysis performed using high throughput sequencing of wild type (DB110) and toxR (TW30) mutant strains of the deep-sea bacterium Photobacterium profundum. ToxR is a transmembrane DNA-binding protein first discovered in Vibrio cholerae and able to regulate numerous genes involved in virulence. In P. profundum the abundance and activity of the same protein is influenced by hydrostatic pressure and is able to regulate genes in a pressure-dependent manner. To better characterize the ToxR regulon, we have compared the genes differentially expressed in response to pressure changes with those whose expression is altered between wild type and toxR mutant strains. Four samples were analyzed: DB110 strain grown at 0.1 MPa, DB110 strain grown at 28 MPa, TW30 strain grown at 0.1 MPa, TW30 strain grown at 28 MPa. Two independent coltures (replicates) were grown for each sample, RNA was extracted from each replicate and RNAs from the two replicates were pooled together to reduce biological variability. No replicates were included in experimental design.
Project description:Genomic DNA extracted from two different Photobacterium profundum strains: SS9 strain (completely sequenced and used to made the microarray) and DSJ4 strain were labeled with Cy3 and Cy5 fluorophores and competitively hybridized on the microarray built on the basis of the SS9 strain genomic sequence. Aim: the identification of the genomic regions absent in the DSJ4 strain with respect to the SS9 strain. The SS9 strain was isolated from the Sulu Trench and display an optimum growth at 28 MPa (2800 metres of depth). The DSJ4 strain was recovered from a sediment sample obtained from the Ryukyu Trench (Japan) at a depth of 5110 m and displays an optimum growth at 10 MPa (but shows no significant change in growth at pressure up to 50 MPa).
Project description:P. profundum SS9 strain cells were grown at two different pressure conditions 45 MPa and at 28 MPa. RNA extracted from the two different cultures was labelled with Cy5 and Cy3 and competitively hybridized on the same slide.
