ABSTRACT
The cell cycle of donor cells as a major factor that affects cloning efficiency remains debatable. G2/M phase cells as a donor can successfully produce cloned animals, but a minimal amount is known regarding nuclear remodeling events. In this study, porcine fetal fibroblasts (PFFs) were carefully synchronized at G1 or M phase as donor cells. Most of the cloned embryos reconstructed from PFFs at G1 (G1-embryos) or M (M-embryos) phase formed a pronucleus-like nucleus (PN) within 6-h post fusion (hpf), but the M-embryos formed PN earlier than the G1-embryos did. Moreover, 77.4% of the M-embryos formed two PNs, whereas the G1-embryos formed a single PN. The rate of extrusion of polar body-like structures by the M-embryos was significantly lower than that extruded by the G1-embryos (26.3% vs. 37.1%, P < 0.05), and DNA synthesis in most embryos in both groups was initiated at 9-12 hpf. Most of the M-embryos were octoploid before the first cleavage. Furthermore, 81.25% of the blastomeres of blastocysts developed from the M-embryos showed abnormal ploidy compared with those developed from the G1-embryos (22.55%). However, some of the blastomeres remained diploid in all the M-embryos tested. A portion of the blastomeres restored normal diploidy in some of the M-embryos at the blastocyst stage. This finding provides an explanation for M-embryos developing to term.
Subject(s)
Blastocyst/cytology , Cloning, Organism/methods , Diploidy , Fibroblasts/cytology , Tetraploidy , Animals , Cell Cycle/genetics , Cellular Reprogramming/genetics , Embryo, Mammalian , Karyotype , SwineABSTRACT
mtDNAs of T type wheat cytoplasmic male sterile lines 75-3369A and their maintainers 75-3369B were isolated and digested completely with restriction endonuclease BamH I, EcoR I, Hind III. The mitochondrial genomic difference between CMS line and maintainers were revealed by Southern hybridization with probes of atp6 from Oenothera and nad3/rps12, cox I from wheat mitochondria. Southern hybridization showed the differences between these two lines. Discussion was made on the change of the DNA composition and the male sterile cytoplasm.
Subject(s)
DNA, Mitochondrial/genetics , Triticum/genetics , Blotting, Southern , FertilityABSTRACT
Adenosine and its derivatives 1 nmol.L-1 -0.1 mmol.L-1 elicited negative chronotropic effect on cultured rat myocardial cells. The respective IC50 were: L-PIA 85 +/- 10 greater than NECA 160 +/- 64 greater than adenosine 361 +/- 41 nmol.L-1. The negative chronotropic effect was antagonized by IBMX. [3H]Adenosine bound to myocardial cells of rat in a saturable way with a Kd of 238 nmol.L-1 and a Bmax of 7.6 pmol/10(8) cells. The binding of [3H]adenosine was competitively antagonized by adenosine, L-PIA, NECA, and IBMX. These results indicated the existence of adenosine receptor in rat myocardial cells.
