High-throughput telomere length measurement at nucleotide resolution using the PacBio high fidelity sequencing platform

Abstract

Telomeres are specialized nucleoprotein structures at the ends of linear chromosomes. The progressive shortening of steady-state telomere length in normal human somatic cells is a promising biomarker for age-associated diseases. However, there remain substantial challenges in quantifying telomere length due to the lack of high-throughput method with nucleotide resolution for individual telomere. Here, we describe a workflow to capture telomeres using newly designed telobaits in human culture cell lines as well as clinical patient samples and measure their length accurately at nucleotide resolution using single-molecule real-time (SMRT) sequencing. Our results also reveal the extreme heterogeneity of telomeric variant sequences (TVSs) that are dispersed throughout the telomere repeat region. The presence of TVSs disrupts the continuity of the canonical (5’-TTAGGG-3’)n telomere repeats, which affects the binding of shelterin complexes at the chromosomal ends and telomere protection. These findings may have profound implications in human aging and diseases.

Biomarkers of aging

“…Recently, SMRT sequencing developed a high throughput TL measurement at nucleotide resolution using the PacBio high fidelity (HiFi) sequencing platform on purified genomic DNA containing telomeric sequence after hybridizing and ligating the telomeric single-stranded G-rich overhangs at the 3′ ends of chromosomes (Tham et al, 2023). This assay not only provides a high-throughput and accurate assay to detect TL at a high resolution but also reveals the presence of telomeric variant sequences (TVSs) interspersed within the long tracts of canonical telomeric repeat regions, which will provide more information for the clinic and research of telomere biology.…”

The associations of socioeconomic status with incident dementia and Alzheimer’s disease are modified by leucocyte telomere length: a population-based cohort study

“…Risk factors such as SES may act as important considerations for community screening of neurodegenerative diseases as well as designing of tailored intervention programs to prevent against cognitive impairment. Further research is needed to improve technologies for measuring biological age (telomere length) at-scale, accuracy and cost effectiveness 57 , 58 .…”

Impact of DNA Extraction Methods on Quantitative PCR Telomere Length Assay Precision in Human Saliva Samples

“…Some new approaches involve whole genome sequencing followed by analysis of the repetitive telomeric sequences (Lee et.al., 2017). Nanopore technology, in which long sequencing reads enable the measurement of the entire telomere sequence is especially promising (Fang et al, 2022;Tham et al, 2023). Perhaps, with further characterization, high-throughput qPCR TL results could be reliably and accurately validated.…”