TeloTool: a new tool for telomere length measurement from terminal restriction fragment analysis with improved probe intensity correction

Abstract

Telomeres comprise the protective caps of natural chromosome ends and function in the suppression of DNA damage signaling and cellular senescence. Therefore, techniques used to determine telomere length are important in a number of studies, ranging from those investigating telomeric structure to effects on human disease. Terminal restriction fragment (TRF) analysis has for a long time shown to be one of the most accurate methods for quantification of absolute telomere length and range from a number of species. As this technique centers on standard Southern blotting, telomeric DNA is observed on resulting autoradiograms as a heterogeneous smear. Methods to accurately determine telomere length from telomeric smears have proven problematic, and no reliable technique has been suggested to obtain mean telomere length values. Here, we present TeloTool, a new program allowing thorough statistical analysis of TRF data. Using this new method, a number of methodical biases are removed from previously stated techniques, including assumptions based on probe intensity corrections. This program provides a standardized mean for quick and reliable extraction of quantitative data from TRF autoradiograms; its wide application will allow accurate comparison between datasets generated in different laboratories.

Protection of Arabidopsis Blunt-Ended Telomeres Is Mediated by a Physical Association with the Ku Heterodimer

“…However, despite its severely reduced ability to bind DNA, the At18 complex associated to telomeric chromatin as well as wild-type or At13 Ku complexes. While this result is consistent with the observed proficiency of At18 (Göhring et al, 2014). Error bars represent SD from at least three independent samples.…”

DDM1 guards against telomere truncation in Arabidopsis

“…1b, 2). Mean telomere length (mean TRF) was measured using the TeloTool (Gohring et al 2014). The average bulk telomere length for at least five individual plants (biological replicates) from each generation was calculated.…”

“…Terminal restriction fragment (TRF) analysis was performed using 50 μg genomic DNA (gDNA) digested with Tru1l , resolved on an 0.8% agarose gel and hybridized with [ 32 P] 5′ end-labeled (T 3 AG 3 ) 4 probe (Fitzgerald et al 1999). The TeloTool was used for TRF quantification (Gohring et al 2014). The mean TRF, standard deviation, the range, the max signal intensity were calculated using the TRF profile of individual plants.…”

A balance between elongation and trimming regulates telomere stability in stem cells

“…Mean telomere length is shown in each condition calculated with TeloTool software 58 . Quantification of telomere length fold change relative to control samples is shown on the right.…”