Genome-wide transcription start site mapping of Bradyrhizobium japonicum grown free-living or in symbiosis – a rich resource to identify new transcripts, proteins and to study gene regulationReportar como inadecuado




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BMC Genomics

, 17:302

Prokaryote microbial genomics

Abstract

BackgroundDifferential RNA-sequencing dRNA-seq is indispensable for determination of primary transcriptomes. However, using dRNA-seq data to map transcriptional start sites TSSs and promoters genome-wide is a bioinformatics challenge. We performed dRNA-seq of Bradyrhizobium japonicum USDA 110, the nitrogen-fixing symbiont of soybean, and developed algorithms to map TSSs and promoters.

ResultsA specialized machine learning procedure for TSS recognition allowed us to map 15,923 TSSs: 14,360 in free-living bacteria, 4329 in symbiosis with soybean and 2766 in both conditions. Further, we provide proteomic evidence for 4090 proteins, among them 107 proteins corresponding to new genes and 178 proteins with N-termini different from the existing annotation 72 and 109 of them with TSS support, respectively. Guided by proteomics evidence, previously identified TSSs and TSSs experimentally validated here, we assign a score threshold to flag 14 % of the mapped TSSs as a class of lower confidence. However, this class of lower confidence contains valid TSSs of low-abundant transcripts. Moreover, we developed a de novo algorithm to identify promoter motifs upstream of mapped TSSs, which is publicly available, and found motifs mainly used in symbiosis similar to RpoN-dependent promoters or under both conditions similar to RpoD-dependent promoters. Mapped TSSs and putative promoters, proteomic evidence and updated gene annotation were combined into an annotation file.

ConclusionsThe genome-wide TSS and promoter maps along with the extended genome annotation of B. japonicum represent a valuable resource for future systems biology studies and for detailed analyses of individual non-coding transcripts and ORFs. Our data will also provide new insights into bacterial gene regulation during the agriculturally important symbiosis between rhizobia and legumes.

KeywordsBradyrhizobium Nodule RNA-seq Transcription start site Promoter prediction Proteogenomics Genome re-annotation Antisense RNA Internal transcription start site AbbreviationsasRNAantisense RNA

EVCempty vector control strain

Freefree-living state

gbkgene bank sequence format

gffgeneric feature format

IGRintergenic region

kbkilobase

Mbmegabase

Nodnodule

ntnucleotide

PCAprinciple component analysis

qRT-PCRquantitative reverse transcriptase-polymerase chain reaction

rrnribosomal RNA operon

rRNAribosomal RNA

RT-PCRreverse transcriptase-polymerase chain reaction

sRNAsmall RNA

SVMsupport vector machine

TEXterminal exonuclease

TSStranscriptional start site

WTwild type

Electronic supplementary materialThe online version of this article doi:10.1186-s12864-016-2602-9 contains supplementary material, which is available to authorized users.

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Autor: Jelena Čuklina - Julia Hahn - Maxim Imakaev - Ulrich Omasits - Konrad U. Förstner - Nikolay Ljubimov - Melanie Goebel -

Fuente: https://link.springer.com/







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