Data from: Extent and variability of interstitial telomeric sequences and their effects on estimates of telomere length

Telomeres often shorten with time, although this varies between tissues, individuals and species, and their length and/or rate of change may reflect fitness and rate of senescence. Measurement of telomeres is increasingly important to ecologists, yet the relative merits of different methods for estimating telomere length are not clear. In particular the extent to which interstitial telomere sequences (ITSs), telomere repeats located away from chromosomes ends, confound estimates of telomere length is unknown. Here we present a method to estimate the extent of ITS within a species and variation among individuals. We estimated the extent of ITS by comparing the amount of label hybridized to in-gel telomere restriction fragments (TRF) before and after the TRFs were denatured. This protocol produced robust and repeatable estimates of the extent of ITS in birds. In five species, the amount of ITS was substantial, ranging from 15% to 40% of total telomeric sequence DNA. In addition, the amount of ITS can vary significantly among individuals within a species. Including ITSs in telomere length calculations always underestimated telomere length because most ITSs are shorter than most telomeres. The magnitude of that error varies with telomere length and is larger for longer telomeres. Estimating telomere length using methods that incorporate ITSs, such as Southern blot TRF and quantitative PCR analyses reduces an investigator's power to detect difference in telomere dynamics between individuals or over time within an individual.端粒（telomeres）通常会随时间推移而缩短，但这一过程因组织类型、个体及物种的不同而存在差异，其长度及/或长度变化速率或可反映物种的适合度与衰老速率。端粒的测量对于生态学家而言愈发重要，但目前尚不清楚不同端粒长度估算方法的相对优劣。尤其值得关注的是，位于染色体末端以外的间质端粒序列（interstitial telomere sequences, ITSs）会在多大程度上干扰端粒长度的估算结果，这一问题仍未明确。本研究提出一种方法，可用于估算某一物种内的ITS占比，以及不同个体间的ITS差异。我们通过比较凝胶内端粒限制性片段（telomere restriction fragments, TRF）在变性前后的标记杂交信号量，来估算ITS的占比。该实验方案可稳定且可重复地得出鸟类样本中ITS占比的估算结果。在五个鸟类物种中，ITS的占比相当可观，范围为总端粒序列DNA的15%至40%。此外，同一物种内不同个体的ITS占比也存在显著差异。将ITS纳入端粒长度计算时，总会低估端粒的实际长度，因为大多数ITS的长度均短于端粒本身。这类误差的幅度随端粒长度变化而改变，且端粒越长，误差幅度越大。采用纳入ITS的端粒长度估算方法（如Southern印迹端粒限制性片段分析法与定量PCR分析），会降低研究者检测个体间端粒动态差异，或是个体内端粒动态随时间变化的效力。