Journal article

Analysis of polyclonal vector integration sites using Nanopore sequencing as a scalable, cost-effective platform

Ping Zhang, Devika Ganesamoorthy, Son Hoang Nguyen, Raymond Au, Lachlan Coin, Siok-Keen Tey

Published : 2019

Abstract

Abstract Vector integration site analysis can be important in the follow-up of patients who received gene-modified cells, but current platforms based on next-generation sequencing are expensive and relatively inaccessible. We analyzed polyclonal T cells transduced by a gammaretroviral vector, SFG.iCasp9.2A.ΔCD19, from a clinical trial. Following restriction enzyme digestion, the unknown flanking genomic sequences were amplified by inverse polymerase chain reaction (PCR) or cassette ligation PCR. Nanopore sequencing could identify thousands of unique integration sites within polyclonal samples, with cassette ligation PCR showing less bias. The assay is scalable and requires minimum capital, w..

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University of Melbourne Researchers