Thu 25th March 2021
Results from developers on human samples — MinION (left) and PromethION (right)
The Ultra-Long DNA Sequencing Kit, which has enabled read lengths of up to 4.2 Mb and maximises the quantity of ultra-long fragments, is now available in store.
In combination with Circulomics Nanobind Kits, the Oxford Nanopore Ultra-Long DNA Sequencing Kit is designed to maximise the quantity of ultra-long reads and has enabled the continuous sequencing of single DNA fragments of up to 3+ Mb externally and 4+ Mb internally. It’s possible for users to achieve more than 100 reads over 1 Mb per PromethION flow cell run using this approach.
The ability to sequence complete ultra-long fragments of DNA with nanopore sequencing provides a totally novel approach to assembly coupled with more comprehensive biological data. Users are able to span large structural variants and highly repetitive regions, whilst gaining information about modifications.
Highly contiguous assemblies
The new kit enables the generation of highly contiguous assemblies — resolving gaps, spanning centromeres and mapping large segmental duplications. Phasing capability is improved, along with mapping of large structural variations and analysis of long repetitive regions.
Users already trialling the kits during a developer release have applied the Ultra-Long Kit to more complete de novo assembly of a range of genomes —from humans to lizards and snakes to cattle. Users have seen 10-20+ Gb of reads per flow cell with an N50 of 50-100+ kb using MinION and 50-100+ Gb of reads with an N50 of 50-100+ kb using PromethION.
‘Whale watching’
Oxford Nanopore devices can sequence DNA/RNA molecules of any length, where the only limit to length is the user’s ability to provide long DNA fragments through the sample preparation steps. The ‘whale scale’ (>1Mb) challenge, initiated by the Long Read Club, took Oxford Nanopore’s transposase based kits and began optimisations to achieve reads over 1Mb long.
Further optimisation of the method, in combination with Circulomics Nanobind CBB Big DNA Kit and Nanobind UL Library Prep Kit, has resulted in the Oxford Nanopore Ultra-Long DNA Sequencing Kit.
Circulomics have released 468 Gb of ultra long data on HG02723 human cell line, including 50X coverage of 100kb+ reads. The data was generated in collaboration with UCSC Genomics Institute. You can access the data here.
Find out more about the Ultra-Long DNA Sequencing Kit in the Community.
Buy the Ultra-Long DNA Sequencing Kit.
Please note, this kit and extraction from Circulomics maximise your quantity of ultra-long reads significantly but if you have less input material you may want to consider the rapid kit, which will give a few ultra-long reads with intact gDNA.
Glossary
Ultra-long – DNA fragment longer than 100,000 bases in continuous length.
N50 – point at which 50% of your data is contained in reads of that length or longer.
Contiguous assemblies – genome assemblies which can be assembled into fewer pieces (or contigs) than previous attempts, and so contain fewer gaps or misassemblies.