14 Telomere length in cloned camels produced by somatic cell nuclear transfer is not different from that in their naturally produced counterparts

Abstract

There are controversial reports on the restoration of eroded telomere length in offspring produced by somatic cell nuclear transfer (SCNT) in different animal species. To the best of our knowledge, no earlier studies report telomere length in naturally produced or cloned animals in any of the camelid species. Therefore, the present study was conducted to estimate the telomere length in dromedary camels produced by SCNT, and their age-matched naturally produced counterparts by terminal restriction fragment (TRF) analysis. Genomic DNA was extracted from venous blood collected from 6 cloned animals (aged 3, 12, and 24 months), and their age-matched counterparts by a conventional phenol/chloroform protocol. Denatured and neutralized DNA was blotted onto a positively charged nylon membrane and fixed by baking at 80°C for 3h. After washing in a prewarmed digoxigenin (DIG) easy hybridization solution at 42°C for 1h, DNA hybridization was carried out using a telomere (TTAGGG)n-specific, DIG-labelled hybridization probe (Roche Diagnostics, Germany) at 42°C for 4h. Stringent washes were carried out at the same temperature, followed by chemiluminescence reaction. The signals were captured using the Azure Biosystems C600 gel documentation system. Molecular weights of the unknown TRF bands on the gel were calculated using known molecular weight marker provided by Telo TAGGG Telomere Length Assay Kit (Roche Diagnostics). A TeloTool program from MATLAB software with a built-in probe intensity correction algorithm was used for TRF analysis. The experiment was replicated 3 times, and the data, presented as mean±s.e.m., were analysed using a 2-sample t-test (Minitab statistical software). No difference (P>0.05) was found in the mean telomere length of cloned camels compared with their naturally produced age-matched counterparts (21.7±0.3 vs. 22.1±0.3; 21.9±0.3 vs. 22.1±0.3; 22.2±0.5 vs. 22.0±0.1; 20.5±0.5 vs. 22.5±0.7; 20.1±0.1 vs. 22.5±0.7; 21.7±1.1 vs. 22.6±0.2), respectively. In conclusion, this is the first study where telomere length has been reported in naturally produced and cloned dromedary camels produced by SCNT. We found that telomere lengths in cloned camels were similar to those of their age-matched naturally produced counterparts, suggesting that the camel cytoplast reprograms the somatic cell nucleus and restores the telomere length to its totipotency stage.