Article Science Technology

LITE2 takes affordable genome sequencing to a new level

The Earlham Institute's improved Low Input, Transposase Enabled (LITE) protocol is a low-cost method for high-throughput whole genome sequencing with reduced DNA input.

05 February 2025

The need to sequence large numbers of samples has never been greater. Whether exploring biodiversity or the impact of genomic variation on specific traits within populations, researchers rely on genome sequence analyses to understand differences between species and populations at the DNA level.

Ever since the first complete sequence of the human genome was published in the early 2000s, technological advances have made it possible to sequence DNA faster and more cheaply. Yet accessing the technology and expertise can be a limiting factor for research groups and SMEs.

Moreover, sequencing thousands of genomes remains expensive and time consuming, mainly due to the cost of processing steps prior to sequencing, known as library preparation.

The original LITE protocol, developed at the Earlham Institute several years ago, can be used to sequence any type of organism - from microbes to plants and animals.

The cost savings result from combining miniaturisation and robotics to reduce the reaction volume - shrinking reagent quantities - and labour.

“We can process a large number of samples, with minimal input and without compromising on the quality of the resulting data,” says Dr Karim Gharbi, Head of the Institute’s Technical Genomics Group.

Compared with the standard manufacturers’ library preparation kits, which often require between 100 and 500 nanograms of DNA, the LITE protocol works with as little as 15 nanograms.

By making small but important changes to the workflow, the team have managed to make significant improvements and are now excited to present LITE2.

TECAN automated liquid handling platform in the sequencing labs at the Earlham Institute

Reduced bias and increased throughput

A key advantage of the LITE2 protocol is that it uses a new transposase enzyme that is more tolerant of DNA stretches with a very high percentage of guanine (G) and cytosine (C) bases.

GC-rich sequences are very stable and difficult to amplify, leading to biased genome coverage.

“With the LITE2 transposase we can achieve a more even coverage of the genome, and confidently use the protocol to sequence species known to have high GC content,” says Thomas Brabbs, Senior Scientist in the Technical Genomics Group.

When the team tested the protocol on samples of Pseudomonas aeruginosa, which has a genome with over 66 per cent GC content, the dropout rate because of high GC was only 3 per cent with LITE2 compared with 35 per cent using the original version.

Another important improvement in the LITE2 protocol is the increased number of samples that can be sequenced together, also known as multiplexing.

“By using a much-expanded set of robust barcodes, we can pool more samples in the sequencing lane, making better use of our capacity and further lowering the cost per sample,” Thomas adds.

I've worked closely with the Technical Genomics group and used the LITE2 protocol to extend our budget to sequence a larger number of samples and capture more genetic diversity.

Charles Dickinson, PhD researcher

Daniella Shipley, Research Assistant, using the TECAN robotic liquid handling platform

Daniella Shipley, Research Assistant, using the TECAN robotic liquid handling platform

Smaller samples, bigger discoveries

As well as population-scale studies, the LITE2 protocol is particularly well suited to microbiome or metagenomic analysis as its reduced bias means that the data more accurately reflect the species composition of the sample.

The Technical Genomics team has already shown in mock metagenomic samples that the proportion of sequencing data from each species closely matches the DNA input, and is applying the improved protocol to sequence microbiomes in soil and the human gut.

The protocol’s low cost makes it economically feasible to perform very large population studies or to enable smaller projects with a modest budget.

A growing number of studies are seeking to characterise natural genetic diversity within a species in relation to particular traits. Plant breeders, for example, are keen to find genetic variations associated with increased yield under changing climate conditions or resistance to disease.

Charles Dickinson is a CASE PhD student at the Earlham Institute, collaborating with Elsoms Seeds Ltd. His project is using genomic resources for commercial herb crops to investigate the genetic differences that drive phenotypic variation in traits associated with flowering.

“Lowering per-sample costs is central to realising the potential of genomics in herb crop breeding,” says Charles.

“I've worked closely with the Technical Genomics group and used the LITE2 protocol to extend our budget to sequence a larger number of samples and capture more genetic diversity.

“This has increased the power of our experiments to reveal the genetics behind complex traits and allowed us to explore global variation in herb crops in greater detail than before.”

The protocol is also particularly appealing to researchers working with very small samples because it requires minimal amounts of DNA. This makes it ideal for any study where collecting DNA isa challenge, such as when working with endangered species or small organisms.

By using a much-expanded set of robust barcodes, we can pool more samples in the sequencing lane, making better use of our capacity and further lowering the cost per sample.

Thomas Brabbs, Senior Scientist

Tom Brabbs, Senior Research Assistant in the Genomics Group at EI

Senior Scientist Tom Brabbs, using the TECAN robotic liquid handling platform

Try LITE

The LITE2 protocol is available for both academic and industry users through Transformative Genomics - the Earlham Institute’s BBSRC-supported National Bioscience Research Infrastructure.

Transformative Genomics brings together the Institute’s technical expertise, equipment, and facilities to both engage with and exploit emerging technologies for genome science.

Its three particular areas of focus are long-read sequencing, single-cell and spatial analysis, and genomics at scale - something which LITE2 enables.

Experienced project managers in the group can offer expert advice on every step of the sequencing process, as well as bioinformatics services also available on site.

“LITE2 really shines when coupled with ultra-high-throughput sequencing on the Illumina NovaSeq X,” Karim says.

“This has brought affordable whole-genome sequencing to a new level.”

Authored by Monica Hoyos Flight, writing for the Earlham Institute.