Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Time-course RNA-seq based transcriptome of Lactococcus piscium MKFS47

ABSTRACT: Lactococcus piscium strain MKFS47 is a psychrotrophic spoilage lactic acid bacterium, isolated from the cold-stored modified atmosphere packaged broiler filet strips with the first signs of spoilage. For the experiment L. piscium MKFS47 was grown in MRS broth without acetate with 2% glucose, samples were taken at 3h, 5h and 11h in three replicates. The extracted RNA was sequenced using SOLiD 5500XL. RNA-seq reads were mapped against L. piscium MKFS47 genome and were counted per gene using Lifescope software. The experiment was conducted to identify the time-course differential expression of the L. piscium MKFS47 genes.

INSTRUMENT(S): AB SOLiD 4 System

ORGANISM(S): Lactococcus piscium

SUBMITTER: Margarita Andreevskaya

PROVIDER: E-MTAB-3245 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Mostly, lactic acid bacteria (LAB), including food-spoilage-associated, grow in communities consisting of several microbial species. The interspecies interactions eventually shape the structure and global activity of a given microbial community. Generally, the knowledge on system level responses of LAB (especially food-spoilage-associated) during such interactions is very limited. To study transcriptome responses during interactions between three MAP meat-spoilage-associated LAB (Leuconostoc gelidum subsp. gasicomitatum LMG 18811T, Lactococcus piscium MKFS47 and Lactobacillus oligofermentans LMG 22743T) we grew them separately in individual cultures and in mixed cultures pairwise (three combinations) and all together (triple culture) in three replicates on a glucose-containing growth medium (MRS) under microaerobic conditions at 25 C, samples were taken at three time points (3, 5 and 11 h) and extracted RNA were sequenced. The experiments were performed in two batches. At first (batch 1), co-cultivation of Le. gelidum and Lc. piscium accompanied with their individual cultures was performed and processed. The raw RNA-seq data for the individual culture of Lc. piscium from the batch 1 were uploaded earlier and are available in the ArrayExpress database under accession number E-MTAB-3245. Later (batch 2), two other pairwise cultures (Le. gelidum + Lb. oligofermentans and Lc. piscium + Lb. oligofermentans) and the triple culture were grown together with the individual cultures of all three LAB. Designations used for the sample names: G: Le. gelidum; P: Lc. piscium; O: Lb. oligofermentans; GO, PO, PG: pairwise cultures of the corresponding species; OPG: triple culture; b1: batch 1; b2: batch 2. Example: 3G2_b1: 3 h, Le. gelidum, 2nd replicate, batch 1; 11PO3_b2: 11 h, pairwise culture of Lc. piscium and Lb. oligofermentans, 3d replicate, batch 2. One sample (5PO3_b2) had very low number of reads ~ 9000, and, therefore, was not uploaded under this project. RNA extraction and library construction were done analogously as in the study (Andreevskaya M et al., 2015. Appl. Environ. Microbiol. 81:38003811, doi: 10.1128/AEM.00320-15). Ribosomal RNA was omitted. Libraries were sequenced in five lanes using SOLiD 5500XL (Life technologies, Foster City, Ca, USA) to produce 75 bp single-end reads. For the data submission, xsq files obtained from SOLiD 5500XL machine, were converted into fastq files. Adapter sequences were removed using cutadapt 1.4.1.

Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goal of this study is to analyse the whole transcriptome of MSCs stimulated with TGFα in order to comprehensively assess the genes regulated by EGFR signalling Methods: Libraries of cDNA were obtained from poly(A)-RNA fractions from not-stimulated or TGFα-stimulated MSCs and sequenced with a SOLiD 5500xl platform. Whole-transcriptome reads were aligned to the version 19 of the human genome (hg19) with the SOLiD LifeScope Genomic Analysis Software version 2.5 (Life Technologies) using the parameters recommended in the user’s manual. The number of observed counts (number of reads/gene) was normalized for the length of the transcript and the number of mapped reads (RPKM) (Reads Per Kilobase per Million of mapped reads). DESeq tool in R package was used to obtain a statistical evaluation of differential gene expression. Results: 1,640 genes resulted highly differentially regulated: 967 genes up-regulated with Fold Induction (FI)≥1.50, and 673 genes down-regulated with FI≤0.50. When highly regulated genes were categorized into enriched categories according to GO molecular function classification and KEGG pathways analysis, a large number of genes coding for potentially secreted proteins or for surface receptors resulted enriched following TGFα treatment. In particular, significant up-regulation of VEGFA, IL6, EREG, HB-EGF, LIF, NGF, NRG1, CCL19, CCL2, CCL25 and CXCL3 was observed. Conclusions: Taken together these findings suggest that EGFR activation in MSCs leads to a significant change in the expression of a wide array of genes coding for secreted proteins that can significantly enhance tumor progression by acting on several mechanisms within the tumor microenvironment mRNA profiles of MSCs starved overnight in serum free medium and treated for 1 hour with recombinant at a concentration of 10 ng/ml by deep sequencing, in quadruplicate, using SOLiD 5500xl.

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Time-course RNA-seq based transcriptome of Lactococcus piscium MKFS47

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