LAUSR

Haploinsufficiency of genes that participate in the telomere elongation and maintenance processes, such as Terc and Tert, often lead to premature aging related diseases such as dyskeratosis congenita and aplastic anemia. Previously we reported that when mouse Terc+/- tail tip fibroblasts (TTFs) were used as the donor cells for somatic cell nuclear transfer (SCNT, also known as "cloning"), the derivative embryonic stem cells (ntESCs) had elongated telomeres. In the present work, we are interested to know if an additional round of SCNT, or recloning, could bring further elongation of the telomeres. Terc+/- TTFs were used to derive the first generation (G1) ntESCs, followed by a second round SCNT using G1-Terc+/- ntESCs as donor cells to derive G2-Tert+/- ntESCs. Multiple lines of G1- and G2-Terc+/- ntESCs were efficiently established, and all expressed major pluripotent markers and supported efficient chondrocyte differentiation in vitro. Comparing to the donor TTFs, telomere lengths of G1-ntESCs were elongated to the level comparable to that in wildtype ntESCs. Interestingly, recloning did not further elongate telomere lengths of the Terc+/- ntESCs. Together, our work demonstrates that while a single round of SCNT is a viable means to reprogram Terc haploinsufficient cells to the ESC state, and to elongate these cells' telomere lengths, a second round of SCNT does not necessarily further elongate the telomeres.