Hi all, I have prior experience in using the Illumina short-read sequencing for bacterial WGS. Given the increasing interest and development on the applications of long-read sequencing technologies (for e.g. Oxford Nanopore and PacBio), I'm looking into the potential of using long-read sequencing.
What I understood from a webinar is, long-read sequencing is more powerful in detecting variants and SNPs, I thought it will be helpful to detect variants, SNPs, and antibiotic resistance genes for my work.
Does anyone have experience in using both technologies? How do you find the pros and cons of both in terms of read quality, final assembly? I assume data processing for long-read sequencing will be more straightforward since you do not have a lot of contigs and gaps? Thanks in advanced for your answer.
Long-read sequencing, or third-generation sequencing, offers a variety of benefits over short-read sequencing. While short-read sequencers generate read of up to 600 bases, long-read sequencing tools usually yield reads in excess of 10 kb. Short-read sequencing is a low-cost, high-accuracy method that is complemented by a multitude of analysis tools and workflows. Natural nucleic acid polymers, on the other hand, can be eight orders of magnitude long, and sequencing them in small amplified segments makes reconstructing and counting the original molecules more difficult. Long reads can help with de novo assembly, mapping certainty, identifying transcript isoforms, and detecting structural variations. Long-read sequencing of native molecules, such as DNA and RNA, also avoids amplification bias while maintaining base modifications. These features, together with ongoing improvements in accuracy, throughput, and cost, have begun to make long-read sequencing a viable choice for a wide range of genomics applications in both model and non-model species.
Thanks Mathew A Beale for your input. I agree that sometimes plasmids and MGEs are hard to resolve using short-read sequencing. Will take this into consideration and see if it aligns with my research questions. Thank you!
Long reads sequencing is great at reconstructing plasmids with great confidence and accuracy and detecting various mobile genetic elements within the genome. It definitely gives you an almost complete genome with great resolution. It's disadvantage is that it is more costly.
Long Reads have a higher error rate then short reads. So, for lowFreg variant calling, short reads are better, due to their very low error rate. Long reads are very helpful to resolve repeat sequences, which are longer then the longest short read you can generate. Here the long continous information shines very bright. Much more powerful is the combination of both technologies. Here you correct the errors in the long reads using short reads from the same sample. This hybrid approach allows the assembly of high quality genomes/transcripts and an very sensitive variant calling with short reads.
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