Figure 3
Reduction in telomere length following siRNA treatment of BEAC cells. SEG-1 cells were transfected with control or telomerase specific siRNAs and telomere length was analyzed either by fluorescent in situ hybridization of metaphase chromosomes, using the Cy3-PNA (C3TA2)3 probe (A, B, C) or by southern hybridization of genomic DNA to a telomere-specific probe (D). (A) SEG-1 cells were treated with control siRNAs for 3 weeks and telomeres on individual chromosomes were analyzed using Telomere-FISH, as described. Telomeres shown as red dots can be seen on most, if not all chromosomes. (B) SEG-1 cells were treated with telomerase specific (Tel) siRNAs for 3 weeks and telomeres were analyzed using Telomere-FISH. Majority of chromosomes are without any telomeres (absence of red dots). The chromosomes which have telomeres (red dots), have a relatively weaker signal, indicative of shorter telomeres; (C) Bar graph showing number of telomeres (red dots) found per 5 cells analyzed at random. (D) SEG-1 cells, transfected with control (Cont) or telomerase specific (Tel) siRNAs, were cultured for 3 weeks. Genomic DNA was isolated and median telomere length was determined by southern hybridization, as described. Median Telomere Length (TL) for each treatment is indicated by arrows.