RNAseq and Methylseq

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Biotechvana is pleased to offer data analysis services for comparative studies to test differential expression of mRNA and non-coding RNA (both long and small) between different RNAseq samples and conditions. We also offer analytical services for epigenetic studies to test differential methylation of gene features and regulatory elements(CpG islands, promoters, introns, exons) between Methylseq samples o similar (e.g. inmmuno-precipitation). Although each study needs to be personalized depending on your experiment design and the species to be resequenced, below we summarize the usual workflow followed by us in these types of studies.

RNAseq Studies

For mRNA and Long ncRNA

1) Preprocessing of raw data

• Quality analysis of raw sequence data files (single or pair-end)
• Preprocessing of reads (either single or pair-end) including demultiplex and removing of low-quality sequences, primers/adapter remnants
  and artifacts and (when appropriate) trimming/clipping, of reads.

2) Mapping on reference

• Reference indexation
• Fastq mapping on the reference genome/transcriptome
• Inference of mapping metrics

3) Quantification and normalization

• Junction Libraries creation
• Transcriptome assembly and summarization of genes and/or isoforms
• Normalization

4) Testing Differential Expression

• Testing differential expression of genes, exons, isoforms, promoters between samples
• Differential enrichment analysis of gene ontology categories (GO-seq)
• Differential enrichment analysis of metabolic pathways

5) Downstream analyses

• Data integration and interrogation
• Protein-Protein interaction (and other) networks
• Correlation analyses
• Other statistical analyses

For Small and micro RNAs

1) Preprocessing of raw data

• Quality analysis of raw sequence data files (either single or pair-end)
• Preprocessing of reads including demultiplex and removing of low-quality sequences, primers/adapter remnants and artifacts and (when
  appropriate) trimming/clipping, of reads.

2) Mapping on reference

• Fastq mapping on the reference genome/transcriptome
• Inference of mapping metrics

3) Quantification and normalization

• Establishment of polarity and sRNA size
• Determination of loci and target genes
• Determination of loci nd target genes characteristics: intergenic or exon regions
• Normalization within and between samples

4) Testing Differential Expression

• Testing differential expression of sRNAs between samples
• Characterization and functional analysis of loci and target genes

5) Downstream analyses

• Identification of phasing sRNAs
• Identification of hairpin structure prediction for miRNAs
• Data integration and functional analysis of loci and target genes
• Characterization of miRNA polymorphisms
• Correlation analyses
• Other statistical analyses

Methylseq and other epigenetic studies

1) Preprocessing of raw data

• Quality analysis of raw sequence data files (single or pair-end)
• Preprocessing of reads (either single or pair-end) including demultiplex and removing of low-quality sequences, primers/adapter remnants
  and artifacts and (when appropriate) trimming/clipping, of reads.

2) Mapping on reference

• Genome methylation reference reconstruction
• Fastq mapping on the reference genome
• Inference of mapping metrics

3) Testing differential methylation

• Testing differential methylation of genomic regions between samples and conditions
• Annotation of regulatory and gene features (CpG islands, promoters, intergenic regions, exons, introns etc) contained by, or overlapping with methylated regions
• Differential enrichment analysis of gene ontology categories (GO-seq)
• Differential enrichment analysis of metabolic pathways

4) Downstream analyses

• Testing differential methylation of genomic regions between samples and conditions
• Annotation of regulatory and gene features (CpG islands, promoters, intergenic regions, exons, introns etc) contained by, or overlapping with methylated regions
• Differential enrichment analysis of gene ontology categories (GO-seq)
• Differential enrichment analysis of metabolic pathways

If you are interested in getting more information about our services for analysis of RNAseq or Methylseq data, please contact us at biotechvana(at)biotechvana.com and we will be glad to pass you on a quote or organize a personalized meeting with you in order to discuss and evaluate your experiment.