title: Comparison of two approaches to nuclear transfer in the bovine: hand-made cloning with modifications and the conventional nuclear transfer technique.
abstract: The aim of the present study was to compare the in vitro and in vivo developmental competence of hand-made cloning (HMC) embryos with the conventional nuclear transfer (NT) method using five somatic cell lines and in vitro-fertilised (IVF; control) embryos. Modifications to the HMC procedure included fusion efficiency optimisation, effect of cytoplasmic volume and cloned embryo aggregation. The developmental competence of blastocysts from each of the treatment groups and cell lines used was assessed following transfer to 345 recipients. Vitrification was also used to enable management of recipient resources and to assess the susceptibility of membranes to cryopreservation following zona removal. Increasing cytoplasmic volume to 150% or aggregating two embryos improved the blastocyst development rate and increased the total cell number. Although HMC embryo transfers established a significantly higher pregnancy rate on Day 30 than fresh IVF or NT embryo transfers, the overall outcome in terms of cloned live births derived from either fresh or vitrified/thawed HMC or NT embryo transfers across the five cell lines did not differ. The birth and continued survival of clones produced with HMC technology with equivalent efficiency to NT shows that it can be used as an alternative method for the generation of cloned offspring in the bovine.
authors.affiliation: Centre for Early Human Development, Monash Institute of Medical Research, Monash University, Clayton, Victoria, Australia. tayfur.tecirlioglu@med.monash.edu.au
authors.forename: R Tayfur
authors.initials: RT
authors.lastname: Tecirlioglu
authors.forename: Melissa A
authors.initials: MA
authors.lastname: Cooney
authors.forename: Ian M
authors.initials: IM
authors.lastname: Lewis
authors.forename: Natasha A
authors.initials: NA
authors.lastname: Korfiatis
authors.forename: Renee
authors.initials: R
authors.lastname: Hodgson
authors.forename: Nancy T
authors.initials: NT
authors.lastname: Ruddock
authors.forename: Gábor
authors.initials: G
authors.lastname: Vajta
authors.forename: Shara
authors.initials: S
authors.lastname: Downie
authors.forename: Alan O
authors.initials: AO
authors.lastname: Trounson
authors.forename: Michael K
authors.initials: MK
authors.lastname: Holland
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: French
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000502:physiology; D002417:Cattle / Q000196:embryology; D019976:Cloning, Organism / Q000379:methods / Q000662:veterinary*; D015925:Cryopreservation / Q000662:veterinary; D003571:Cytochalasin B / Q000494:pharmacology; D003593:Cytoplasm / Q000648:ultrastructure; D046149:Embryo Culture Techniques / Q000662:veterinary; D004624:Embryo Transfer / Q000662:veterinary; D047108:Embryonic Development; D005260:Female; D005307:Fertilization in Vitro / Q000662:veterinary; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000187:drug effects / Q000648:ultrastructure; D011247:Pregnancy; D012710:Serum Albumin, Bovine / Q000494:pharmacology
chemical_list: D012710:Serum Albumin, Bovine; D003571:Cytochalasin B