Comparative potential of cultured skin fibroblast, cumulus, and granulosa cell to produce somatic cell nuclear transfer (SCNT) preimplantation embryos in buffaloes (Bubalus bubalis) in relation to gene expressions.
To improve the efficiency of somatic cell nuclear transfer (SCNT)-derived embryos in buffaloes (Bubalus bubalis), skin fibroblast, cumulus, and granulosa cells were cultured up to the 15th passage and cloned embryos were produced from each cell type. At the 15th passage the cumulative population doublings (CPDs) in cumulus cells was higher (60.78) than skin fibroblast (57.12) and granulosa (56.05) cell lines. Gene expression of chromatin remodelling proteins, that is, HDAC1, DNMT1, DNMT3a, and DNMT3b, were comparable at all five passages (P-3, P-6, P-9, P-12, and P-15) groups in cumulus cells but different in skin fibroblast and granulosa cells. Cleavage and blastocyst production rate in cumulus (65.9 and 27.4%)-derived embryos was higher than skin fibroblast (63.8 and 24.3%) and granulosa (62.5 and 22.3%)-derived embryos. Expressions of HDAC1, DNMT1, and DNMT3a mRNA in cumulus-derived blastocysts were similar to IVF blastocysts (control), whereas skin fibroblast and granulosa-derived blastocysts expression was significantly different (p < or = 0.05). DNMT3b mRNA expression in all the three donor cell types and IVF control were similar. The expression pattern of these genes showed the effect of donor cell type with different epigenetic reprogramming capabilities for SCNT embryo production rate. Overall, results indicated that cumulus cells are the best nuclear donor for SCNT.
Pandey
Alok
A
Transgenic Research Laboratory, National Bureau of Animal Genetic Resources, Karnal, India.
Gupta
S C
SC
Gupta
Neelam
N
D000818:Animals; D001483:Base Sequence; D001755:Blastocyst; D002020:Buffaloes / Q000235:genetics*; D049109:Cell Proliferation; D002478:Cells, Cultured; D017931:DNA Primers; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D015870:Gene Expression; D006107:Granulosa Cells / Q000166:cytology*; D053652:Nuclear Transfer Techniques; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012867:Skin / Q000166:cytology*
D017931:DNA Primers