Establishment of a Conditional Transgenic System Using the 2A Peptide in the Female Mouse Germline

Transgenic mice are essential research tools in developmental biology studies. The 2A peptide allows multiple genes to be expressed simultaneously at comparable levels in somatic cells, but there are no reports of it being used successfully in germ cells. We constructed a Cre/loxP-based conditional vector containing the 2A peptide to significantly enhance the expression of a reporter and target gene from a constitutive promoter in oocytes. Mice with a transgene insertion containing the chicken β-actin promoter, floxed EGFP-polyA cassette, mCherry reporter, 2A peptide and target gene DNA methyltransferase 3A2 (Dnmt3a2) were crossed with TNAP- or Vasa-Cre mice to produce offspring, in which mCherry and DNMT3A2 proteins were highly expressed in oocytes upon Cre-mediated removal of EGFP-polyA. This novel transgenic mouse line based on the 2A expression system can serve as a useful tool for examining gene function during oogenesis.     

Thinning alters crown dynamics and biomass increment within aboveground tissues in young stands of Chamaecyparis obtusa

The stand density of a forest affects the vertical distribution of foliage. Understanding the dynamics of this response is important for the study of crown structure and function, carbon-budget estimation, and forest management. We investigated the effect of tree density on the vertical distribution of foliage, branch, and stem growth, and ratio of biomass increment in aboveground tissues; by monitoring all first-order branches of five trees each from thinned and unthinned control stands of 10-year-old Chamaecyparis obtusa for four consecutive years. In the control stand, the foliage crown shifted upward with height growth but the foliage quantity of the whole crown did not increase. In addition, the vertical distribution of leaf mass shifted from lower-crown skewed to upper-crown skewed. In the thinned stand in contrast, the foliage quantity of individual crowns increased two-fold within 4 years, while the vertical distribution of leaf mass remained lower-crown skewed. The two stands had similar production rates, numbers of first-order branches per unit of tree height, and total lengths of first-order branches. However, the mortality rate of first-order branches and self-pruning within a first-order branch were significantly higher in the control stand than in the thinned stand, which resulted in a higher ratio of biomass increment in branch. Thinning induced a higher ratio of biomass increment in foliage and lower in branch. The increased foliage quantity and variation in ratio of biomass increment after thinning stimulated stem growth of residual trees. These results provide information that will be useful when considering thinning regimes and stand management.     

Effect of improvement cutting on the community structure of aphyllophoraceous fungi on Okinawa Island

Improvement cutting, which decreases tree density by cutting and thinning while leaving logging residues, has been practiced on Okinawa Island since the 1970s and changes not only the vegetation composition but also the amount and composition of woody debris on the forest floor. To quantitatively compare the effect of improvement cutting with the effects of woody debris and vegetation on the community structure of aphyllophoraceous fungi, we conducted a field survey of vegetation composition, the amount and composition of woody debris, and the aphyllophoraceous fungi flora on Okinawa Island in December 2006. We established 3 plots in forest stands without logging and 13 plots in a forest that had undergone improvement cutting from 1 to 22 years before our field survey. The number and volume of woody debris tended to be highest soon after improvement cutting. We recorded 1,172 fungal samples belonging to 81 species in four orders. Fungal species richness was correlated with the composition of the woody debris. Fungal species composition in forest plots 1 year after improvement cutting differed from most of the other logged forests. Variation partitioning revealed that the effects of improvement cutting explained 14.1 % of the total variation in fungal species composition, and that differences in woody debris and vegetation not related to improvement cutting explained 40.4 %. Changes in the composition of woody debris caused by improvement cutting explained at most 2.7 % of the variation. This suggests that improvement cutting does not have a strong effect on fungal community structure, but woody debris does.     

The sex determining gene of medaka: a Y-specific DM domain gene (DMY) is required for male development

Although the sex-determining gene Sry has been identified in mammals, no comparable genes have been found in non-mammalian vertebrates. To clone positionally the sex-determining region of the medaka, Oryzias latipes, we generated a Y congenic strain to highlight the genetic differences between the X and Y chromosomes from inbred strains of medaka. We used recombinant breakpoint analysis and deletion analysis of the Y chromosome of a congenic XY female to restrict the sex-determining region to 250-kb stretch of the Y chromosome. Shotgun sequencing of this region predicted 27 genes. Three of these genes were expressed during sexual differentiation. However, only one gene was Y specific. The full-length cDNA sequence of this gene encodes a putative protein of 267 amino acids, including the highly conserved DM domain. We thus named it DMY. To establish a role for DMY during sexual differentiation, we screened wild medaka populations for naturally occurring DMY mutants. Two XY females with distinct mutations in DMY were found in separate populations. The first heritable mutant – a single insertion in exon 3 and the subsequent truncation of DMY – resulted in all XY female offspring. Similarly, the second XY mutant female showed reduced DMY expression with a high proportion of XY female offspring. Furthermore, during normal development, DMY is expressed only in somatic cells of XY gonads. These findings strongly suggest that the sex-specific DMY is required for normal testicular development and is a prime candidate for the medaka sex-determining gene.     

The association between gall bladder mucoceles and hyperlipidaemia in dogs: A retrospective case control study

The diagnosis of gall bladder mucoceles (GM) in dogs has become increasingly frequent in veterinary medicine. Primary breed-specific hyperlipidaemia is reported in Shetland Sheepdogs and Miniature Schnauzers, breeds in which GM are known to occur more frequently than in other breeds. The objective of this study was to evaluate the association between GM and hyperlipidaemia in dogs. The study design was a retrospective case control study. Medical records of dogs diagnosed with GM at the Veterinary Medical Centre of The University of Tokyo between 1 April 2007 and 31 March 2012, were reviewed.Fifty-eight dogs with GM and a record of either serum cholesterol, triglyceride, or glucose concentrations were included in the study. Hypercholesterolaemia (15/37 cases; odds ratio [OR]: 2.92; 95% confidence interval [CI]: 1.02–8.36) and hypertriglyceridaemia (13/24 cases; OR: 3.55; 95% CI:1.12–15.91) showed significant association with GM. Pomeranians (OR: 10.69), American Cocker Spaniels (OR: 8.94), Shetland Sheepdogs (OR: 6.21), Miniature Schnauzers (OR: 5.23), and Chihuahuas (OR: 3.06) were significantly predisposed to GM. Thirty-nine out of 58 cases had at least one concurrent disease, including pancreatitis (five cases), hyperadrenocorticism (two cases), and hypothyroidism (two cases). A significant association between GM and hyperlipidaemia was confirmed, suggesting that hyperlipidaemia may play a role in the pathogenesis of GM.