Canine mdr1 gene mutation in Japan

Frequency of the 4-bp deletion mutant in canine mdr1 gene was examined in 193 dogs of eight breeds in Japan. The mutant allele was found in Collies, Australian Shepherds, and Shetland Sheepdogs, where its respective frequencies were 58.3%, 33.3%, and 1.2%. The MDR1 protein was detected on peripheral blood mononuclear cells (PBMC) from a MDR1/MDR1 dog, but not on PBMC from a mdr1-1Delta/mdr1-1Delta Collie. Rhodamine 123 was extruded from MDR1/MDR1 lymphocytes. That excretion was inhibited by a MDR1 inhibitor, verapamil. On the other hand, Rh123 excretion was not observed from lymphocytes derived from a mdr1-1Delta/mdr1-1Delta Collie. These results indicated that the mutant mdr1 allele also existed in Collie-breed dogs in Japan at high rates and that mdr1-1Delta /mdr1-1Delta dogs have no functional MDR1.     

Cell wall modification during development of mealy texture in the stony-hard peach “Odoroki” treated with propylene

The melting peach cultivar “Akatsuki” [Prunus persica L. Batsh] develops melting texture during ripening, but the stony-hard peach cultivar “Odoroki”, softens and develops a mealy texture when treated with propylene. Cell wall polysaccharides were fractionated as 1,2-diaminocyclohexanetetraacetic acid (CDTA)-, Na₂CO₃-, NaOH-soluble fractions, or the cellulosic residues that remained after solvent extraction. Size-exclusion chromatography of the CDTA-soluble fraction revealed a limited depolymerization of polyuronides in mealy “Odoroki” fruit, accompanying a lower level of endo-polygalacturonase (endo-PG) and a higher level of exo-PG activities compared to melting “Akatsuki” fruit. “Akatsuki” fruit showed increases in the arabinose, galactose and rhamnose contents in the CDTA-soluble fraction. These neutral sugars decreased in the Na₂CO₃-soluble and cellulosic residue fractions during ripening. Similar changes were observed for neutral sugars other than arabinose during softening in mealy “Odoroki” fruit. Arabinose content did not increase in the CDTA-soluble fraction of “Odoroki” fruit; however, arabinose content decreased in the Na₂CO₃-soluble fraction and in the cellulosic residue as observed for “Akatsuki” fruit. A significant increase in α-l-arabinofuranosidase activity, about 10 times higher than for “Akatsuki” fruit, was observed in propylene-treated mealy “Odoroki” fruit. Differences in ripening-associated metabolic changes in the cell walls of these peach cultivars seem to be related to their differences in texture.     

Sequence analysis of VP2 gene of canine parvovirus isolated from domestic dogs in Japan in 1999 and 2000

Seven strains of canine parvovirus (CPV) were isolated from affected dogs in Japan between 1999 and 2000, and their VP2 genes were genetically analyzed. Comparison of the predicted amino acid sequences of VP2 suggested that three field isolates corresponded to CPV type 2a, while the other four to CPV type 2b. The phylogenetic tree constructed from the VP2 genes showed that the newly isolated strains are classified into the cluster consisting of recent Japanese and Taiwanese field isolates, which are distinct from Vietnamese isolates, United States Isolates, or classical CPV type 2. These results suggest that the CPV transmission occurred between Japan and Taiwan in 1990s, and the offspring are still circulating in both countries.     

Application of laser-assisted zona drilling to in vitro fertilization of cryopreserved mouse oocytes with spermatozoa from a subfertile transgenic mouse

Development of assisted reproductive technologies is necessary to obtain fertilized oocytes in a subfertile transgenic mouse strain. Here, we showed the application of laser-assisted drilling of the zona pellucida to in vitro fertilization of cryopreserved mouse oocytes with sperm from subfertile transgenic mice (C57BL/6N-Tg(UCP/FAD2)U8 strain). After cryopreservation by vitrification, the recovery and survival rates of the zona-drilled mouse oocytes were 97% (97/100) and 94% (91/97), respectively. In vitro fertilization of the cryopreserved zona-drilled mouse oocytes with sperm from the subfertile transgenic mice was greatly facilitated (60%, 55/91) compared to that of the cryopreserved zona-intact mouse oocytes (11%, 81/768). In vitro fertilized embryos that developed to the 2-cell stage were again cryopreserved by vitrification, and after warming they were transferred into recipient females. Subsequently, six viable offspring were delivered, and all were confirmed to be transgenic mice. These results indicate that laser-assisted zona drilling of oocytes combined with cryopreservation by vitrification may be a useful approach for large-scale production of in vitro fertilized embryos for managing transgenic mouse strains with reproductive disabilities such as subfertile sperm.     

Twenty-eight new microsatellite loci in chicken and their cross-species amplification in Japanese quail and helmeted guinea fowl

Twenty‐eight original chicken microsatellite markers were isolated and characterized to determine their utility as cross‐reactive markers for comparative genetic mapping in the order Galliformes. Primer pairs were typed in 12 unrelated chickens and also tested on Japanese quail and helmeted guinea fowl deoxyribonucleic acid (DNA). Polymorphism was observed in 23 (82.1%) of the markers and the average number of alleles per locus was 2.9 while the mean heterozygosity was 0.19. Eleven (39.3%) of the chicken markers cross‐reacted with Japanese quail DNA and 2 (7.1%) with helmeted guinea fowl DNA. The cross‐reactive markers described would serve as useful resources for comparative genetic mapping in poultry species belonging to the order Galliformes.     

Relationship between serum leptin concentrations and the marbling scores in Japanese Black Cattle

The objective of this study was to investigate the relationship between marbling and serum leptin concentrations. First, we compared serum leptin concentrations between Japanese Black cattle and Holstein calves, and found that the mean serum concentration was significantly higher (P < 0.05) in Japanese Black cattle (2.67 ± 0.41 ng/mL, n = 13) than in Holstein calves (1.79 ± 0.43 ng/mL, n = 5). We then examined the effects of roughage intake or vitamin A on the serum leptin concentrations in Japanese Black cattle. Although the leptin concentrations for the high roughage group were significantly higher than those for the low roughage group at 4, 7 and 20 months old, there was no significant difference in the carcass characteristics of the two groups. In addition, although there was a significant difference in the marbling score between the high vitamin A group and low vitamin A group, there was no significant difference in serum leptin concentrations between the groups. These results suggest that although serum leptin concentrations were not significantly correlated to the marbling scores they may be related to the genetic advantage for marbling in Japanese Black cattle.