HLA Multiplexed GenDx Amplicons HLA A , B, C, DQB1, and DRB1

Instrument:  PacBio RSII
Chemistry:  C4
Enzyme: P6
RunTime: 180 min

Summary

This dataset includes four SMRT Cells of multiplexed HLA amplicons using GenDx primers for HLA typing, sequenced with P6-C4 chemistry on the PacBio RSII System. Each SMRT Cell contains a 8 different samples with barcodes applied using two different strategies.

• Class I HLA Amplicons: A, B, C
• Class II HLA Amplicons: DQB1, DRB1

More information on GenDx amplicons and the NGSgo reagent kit can be found via the link below. Consensus sequences can be generated with SMRT Analysis Long Amplicon Analysis Protocol version 2.3. Expected amplicon sizes range from 3.1Kb to 4.8Kb. One HLA-A allele drop-out is noted in barcode lbc2--lbc2 from the prototype barcode dataset.

Related Topics

• Amplicon Analysis Documentation
• HLA Getting Started Guide (coming soon)
• PacBio Barcoding Sample Preparation
• PacBio Barcoding Secondary Analysis
• GenDx NGSgo
• GenDX NGSengine

Download Datasets

Adapter-Ligated Barcodes

Symmetric barcodes appended via barcoded SMRTbell adapters to amplicons generated using GenDx NGSgo-AmpX in combination with PacBio library prep reagents.

• SMRT Cell with 8 samples x 5 (Class I/II) amplicons pooled together (7.2Gb)
• 8 samples x 5 (Class I/II) demultiplexed FASTQ raw reads* (267Mb)
• Barcode FASTA
• Expected HLA Types

Prototype Barcode Dataset

Symmetric barcodes appended using a prototype primer kit (not commercially available at time of writing).

• SMRT Cell with 8 samples x 3 (Class I) amplicons pooled together (7.5Gb)
• 8 samples x 3 (Class I) demultiplexed FASTQ raw reads* (338Mb)
• SMRT Cell with 8 samples x 2 (Class II) amplicons pooled together (5.8Gb)
• 8 samples x 2 (Class II) demultiplexed FASTQ raw reads* (254Mb)
• SMRT Cell with 8 samples x 5 (Class I/II) amplicons pooled together (7.1Gb)
• 8 samples x 5 (Class I/II) demultiplexed FASTQ raw reads* (286Mb)
• Barcode FASTA
• Expected HLA Types

* Recommended HLA workflow for PacBio data uses the SMRT Analysis Long Amplicon Analysis (LAA) software to properly handle rich QV metrics contained in PacBio native format. The consensus sequences obtained from LAA can be loaded directly into NGSengine for typing analysis.