mRNA-Seq-8 Sample Prep Kit

The mRNA-Seq 8-Sample Prep Kit is used to build libraries for single-read and paired-end sequencing on all Illumina sequencing systems. Sample preparation is straightforward, using standard molecular biology techniques and requiring minimal hands-on time.

Starting with 1 μg total RNA, mRNA is isolated using polyA selection. The mRNA is then fragmented and randomly primed for reverse transcription followed by second-strand synthesis to create double-stranded cDNA fragments. Ends are repaired with a combination of fill-in reactions and exonuclease activity to produce blunt ends.

An 'A'- base is added to the blunt ends followed by ligation to Illumina Paired-End Sequencing adapters followed by PCR amplification. These adapters contain unique sequencing primer hybridization sites.

Additional sequences complementary to the oligonucleotides in the flow cell are added to the adapter sequences with tailed PCR primers. This is followed by gel-based size selection and purification to create libraries for cluster generation.

Transcriptome Analysis

RNA sequencing has revolutionized the exploration of gene expression. Advances in the sequencing workflow, from sample preparation through data analysis, enable rapid profiling and deep investigation of the transcriptome. With RNA sequencing, you can characterize all transcriptional activity, coding and non-coding, in any organism without a priori assumptions.

Illumina's unique combination of long and short reads, single and paired-end sequencing, strand specificity, and capacity for tens of millions to billions of reads per run allows you to
  • annotate coding SNPs
  • discover transcript isoforms
  • identify regulatory RNAs
  • characterize splice junctions
  • determine the relative abundance of transcripts

With the greatest daily output available for any sequencing system, transcript profiles can be generated in a single day. RNA sequencing reads can be aligned across splice junctions to identify isoforms, novel transcripts and gene fusions. Identify and quantify both rare and common transcripts, with over six orders of magnitude of dynamic range. Reveal the hidden world of non-coding RNA architecture without prior information.

 

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Small RNA Discovery and Analysis

Small RNA sequencing is a powerful application for Illumina Sequencing, enabling the discovery and profiling of microRNAs and other non-coding RNA on any organism, without prior genome annotation. Using low RNA inputs, you can profile the differential expression of known microRNAs as well as detect novel microRNA targets and wide-ranging sequence variation or "iso-miRs" miRBase accessions. With unprecedented sensitivity and dynamic range, Illumina's industry-leading small RNA sequencing methods allow for the most accurate detection and quantification of rare small RNA sequences.

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