Sample Preparation
- Library preparation - Standard Iso-Seq library / Multi- throughput Iso-Seq library/polyA Iso-Seq library
RNA Sequencing (RNA-seq) has become the most frequently used method for the majority of researchers conducting gene expression profiling. However, it is difficult to obtain a complete picture of the transcriptome because short reads cannot accurately assemble complex transcripts.
“Isoform Sequencing” (Iso-seq) developed by Pacific Biosciences (PacBio), is based on long-read sequencing technology. The unique long-read sequencing feature allows this method to identify new isoforms with extraordinary precision. The Iso-Seq application generates full-length cDNA sequences — from the 5’ end of transcripts to the poly-A tail — eliminating the need for transcriptome reconstruction using isoform-inference algorithms. The Iso-Seq method provides accurate information about alternatively spliced exons and transcriptional start sites. It also reveals information about poly-adenylation sites for transcripts across the full complement of isoforms within targeted genes or the entire transcriptome.
We care for your project from the start through to reporting of results. Highly experienced laboratory professionals follow strict quality procedures to ensure the integrity of your results.
Send us a no obligation request for quote and our dedicated sales team will reply to you within 24 hours.
BGI Iso-Seq/PacBio transcriptome sequencing services are executed on the Sequel I/II platform.
We can process your total RNA, whole blood, cell line, fresh frozen tissue samples, with the following general requirements:
Minimum Sample Volume: 15 µl
Recommended
m≥3 µg
c≥300ng/µL
RIN≥8
Required
1µg≤m<3 µg
28S/18S(23S/16S)≥1.4
Besides clean data, BGI offers a range of standard and customized bioinformatics options for your Iso-seq/PacBio transcriptome sequencing. Reports and output data files are delivered in industry-standard file formats: FASTQ, BAM, cout, .xls, .png
Remove the low-quality reads and short reads
Identify the full-length, non-chimeric transcripts and non-full-length, non-chimeric transcripts
Build similarity graph using BLASR, get cluster consensus
Polish the consensus sequences and get high quality full-length, non-chimeric transcripts
Merge consensus sequences of all libraries and remove redundancy
Annotation of the full-length non-chimeric transcripts (Nr、Nt、Swissprot、KEGG、GO、COG and Interpro)
CDS prediction
SSR prediction
Remove the low-quality reads and short reads;
Identify the full-length, non-chimeric transcripts and non-full-length, non-chimeric transcripts
Build similarity graph using BLASR, get cluster consensus
Polish the consensus sequences and get high quality full-length, non-chimeric transcripts
Align to the reference genome with GMAP
Merge consensus sequences of all libraries and remove redundancy
Transcripts classification
Novel transcripts analysis
Long noncoding RNA prediction
Splicing site detection and annotation
Gene fusion detection and annotation
Analysis of Standard Iso-Seq + Gene/Transcripts quantification; differentially expressed gene detection and annotation
Analysis of Standard Iso-Seq + Gene/Transcript quantification; differentially expressed gene detection and annotation; polyA length analysis
Further customization of bioinformatics analysis to suit your needs is available
Please contact your BGI technical representative for details