34 MEIOTIC RECOMBINATION IN SOMATIC CELL NUCLEAR TRANSFER BULLS AND THEIR OFFSPRING

Abstract

In mammals, homologous chromosome pairing and recombination are essential events for meiosis. The generation of reciprocal exchanges of genetic material ensure both genetic diversity and the proper segregation of homologous chromosomes. With the advent of reproductive biotechnologies such as somatic cell nuclear transfer (SCNT) in the livestock industry, these vital steps have begun to be bypassed. Despite this, there have been few studies carried out on the reproductive characteristics of cloned animals and none to date regarding the consequences on the meiotic process. As these procedures grow in popularity and use, the importance of evaluating the long-term viability, health, and productivity of cloned animals and any subsequent offspring in future generations to validate potential applications of the technology is increasingly evident. Previously, studies of recombination and synapsis have focused on the physical observation of chiasmata formation in meiotic chromosomes; however, in recent years, the characterization of proteins that localize to the sites of crossing-over and of proteins present in the synaptonemal complex have permitted the study of meiotic recombination using a precise direct immunocytogenetic approach. Cytological analysis of meiotic cells obtained from the testicular tissue of five normal bulls of proven fertility, two SCNT transgenic bulls (Powell et al. 2004 Biol. Reprod. 71, 210–216, 2004), and four reproductively mature offspring of SCNT bulls was performed in order to detect the effects of SCNT on the meiotic process. Over 50 pachytene cells per animal were analyzed by immunofluorescence using antibodies against the synaptonemal complex protein 3 (SCP3) and the mismatch repair protein 1 (MLH1) located on the crossover sites. Data were analyzed using a mixed model analysis of variance with repeated measurements to determine group effects (SAS 9.1; SAS Institute, Inc., Cary, NC, USA). The average number of crossovers per spermatocyte for the non-SCNT bulls (42 � 4 SD, min: 33, max: 56), SCNT bulls (43 � 5 SD, min: 35, max: 56), and SCNT offspring (46 � 4 SD, min: 37, max: 58) were quite similar among the cells of the same individual; however, inter-individual variation was observed. These results are the first documentation of the normal range of variability of recombination distribution within the cattle genome and suggest that the SCNT process does not affect meiotic recombination. This work was funded by NSERC and the CRC program.