Inbreeding effects on in vitro embryo production traits in Guzerá cattle

Inbreeding has been associated with the impairment of reproductive performance in many cattle breeds. Although the usage of reproductive biotechnologies has been increasing in bovine populations, not much attention has been given to the impact of inbreeding over cow’s performance on artificial reproduction. The objective of this study was to estimate the impact of inbreeding on in vitro embryo production in a Guzerá breed population. The inbreeding coefficient (F), calculated as half of the co-ancestry of the individual’s parents, was used as an estimate of inbreeding. The inbreeding coefficients of the donor, sire (used on in vitro fertilization) and of the embryos were included, separately, in the proposed models either as classificatory or continuous variables (linear and quadratic effects). The percentage of non-inbred individuals (or embryos) and mean F of donors, embryos and sires were 29.38%; 35.76%; 42.86% and 1.98±2.68; 1.32±3.13; 2.08±2.79, respectively. Two different models were considered, one for oocyte production traits and other for embryo production traits. The increase of F of the donor significantly (P<0.05) impaired the number of viable oocytes (N_OV), number of grade I oocytes (N_GI) and number of cleaved embryos (N_CLV). Moreover, the donor’s F influenced the percentage of grade I oocytes (P_GI), percentage of viable embryos (P_EMB) and percentage of cleaved embryos that turned into embryos (P_CXE). No significant (P>0.05) effects were observed for the sire (father of the embryos) inbreeding coefficient over the traits analysed. Embryo’s F influenced (P<0.05) the number of viable embryos (N_EMB), percentage of viable embryos (P_EMB) and percentage of cleaved embryos that turn into embryos (P_CXE). Results suggested that an increase in the inbreeding coefficient might impair the embryos ability to survive through challenges imposed by the in vitro environment. Submitting highly inbred Guzerá female donors to in vitro embryo production may, in the long-term, have negative implications on the number of embryos obtained per cow and increase the relative costs of the improvement programmes based on this technology. High levels of inbreeding should be avoided when selecting Guzerá female donors and planning in vitro fertilization mating.     

Count Bayesian models for genetic analysis of in vitro embryo production traits in Guzerá cattle

Four models for in vitro embryo production traits in Guzerá cattle were compared: Gaussian (untransformed variable – LIN and transformed in logarithmic scale – LOG), Poisson (POI) and zero-inflated Poisson (ZIP). Data consisted of 5716 ovum pick-up and in vitro fertilization records performed in 1205 cows from distinct regions of Brazil. Analyzed count traits were the number of viable oocytes (N_OV), number of grade I oocytes (N_GI), number of degenerated oocytes (N_DG), number of cleaved embryos (N_CLV) and number of viable produced embryos (N_EMB). Heritability varied from 0.17 (LIN) to 0.25 (POI) for N_OV; 0.08 (LOG) to 0.18 (ZIP) for N_GI; 0.12 (LIN) to 0.20 (POI) for N_DG; 0.13 (LIN) to 0.19 (POI) for N_CLV; 0.10 (LIN) to 0.20 (POI) for N_EMB depending on the considered model. The estimated repeatability varied from 0.53 (LOG) to 0.63 (POI) for N_OV; 0.22 (LOG) to 0.39 (ZIP) for N_GI; 0.29 (LIN) to 0.42 (ZIP) for N_DG; 0.42 (LIN) to 0.59 (POI) for N_CLV; 0.36 (LIN) to 0.51 (POI) for N_EMB. The goodness of fit, measured by deviance information criterion and mean squared residuals, suggested superiority of POI and ZIP over Gaussian models. Estimated breeding values (EBV) obtained by different models were highly correlated, varying from 0.92 for N_OV (between LIN-POI) and 0.99 for N_GI (between POI-ZIP). The number of coincident animals on the 10% top EBV showed lower similarities. We recommend POI and ZIP models as the most adequate for genetic analysis of in vitro embryo production traits in Guzerá cattle.     

Embryogenic cells in plant tissue cultures: Occurrence and behavior

Summary Plants develop from meristems where cells proliferate and are partitioned into layers that eventually differentiate to form the various tissues and organs of the plant. A phenomenon unique to certain plant tissues is the ability to inducede nova a range of developmental patterns, including embryogenesis. The temporal and spacial distribution of these developmental competencies suggests that regulatory proteins, rather than a lack of signals or signal receptors, shield specific developmental genes from signals that otherwise would confuse development. Studies involving embryogenic cells demonstrate that they are not strongly shielded from developmental signals, thus they are not determined. Furthermore, the normal development of zygotic and somatic embryos is readily perturbed by abnormal physicochemical environments. This suggests that embryogeny remains developmentally plastic until differentiation is largely completed. The ability to induce somatic embryogenesis from specific tissues by specific signals is providing opportunities to further the molecular characterization of the menagerie of genetic regulation involved in development. Much of this review was presented at a Moet-Hennessy Louis Vuitton sponsored conference, Control of Morphogenesis in Plants, Aix Les Bains, France, September, 1989. Studies in our laboratory were supported in part by grants from the National Aeronautics and Space Administration (Cooperative Agreement No. NCC2-139), the Utah State University Biotechnology Center, and by the Utah Agricultural Experiment Station, Utah State University, Logan, UT 84322-4845 (approved as journal paper no. 3928).     

Characterization of gene expression in double-muscled and normal-muscled bovine embryos

Myostatin, a member of the transforming growth factor-beta superfamily, is a negative regulator of skeletal muscle growth. Cattle with mutations that inactivate myostatin exhibit a remarkable increase in mass of skeletal muscle called double muscling that is accompanied by an equally remarkable decrease in carcass fat. Although a mouse knockout model has been created which results in mice with a 200% increase in skeletal muscle mass, molecular mechanisms whereby myostatin regulates skeletal muscle and fat mass are not fully understood. Using suppressive subtractive hybridization, genes that were differentially expressed in double-muscled vs. normal-muscled cattle embryos were identified. Genetic variation at other loci was minimized by using embryonic samples collected from related Piedmontese x Angus dams or Belgian Blue x Hereford dams bred to a single sire of the same breed composition. Embryos were collected at 31-33 days of gestation, which is 2-4 days after high-level expression of myostatin in the developing bovine embryo. The suppressive subtraction resulted in 30 clones that were potentially differentially expressed, 19 of which were confirmed by macroarray analysis. Several of these genes have biological functions that suggest that they are directly involved in myostatin's regulation of skeletal muscle development. Furthermore, several of these genes map to quantitative trait loci known to interact with variation in the myostatin gene.     

不同启动子对于牛催乳素表达的调控作用

在细胞水平上比较不同启动子对于牛催乳素(bPRL)表达的调控作用.分别构建了以CMV启动子、牛催乳素基因启动子和山羊β-酪蛋白基因启动子作为调控元件的bPRL真核细胞表达载体,分别命名为pCMV、pPRLP和pP1A3.将3种载体分别转染小鼠垂体瘤细胞和小鼠乳腺上皮细胞,使用RT-PCR和定量RT-PCR分析3种启动子启动bPRL在2种细胞系中的表达效果.pCMV在2种细胞中有效表达bPRL;pPRLP在2种细胞中的表达效果与pCMV接近:pP1A3不在垂体细胞中表达,在乳腺细胞中表达.pPlA3具有乳腺表达特异性;pPRLP能够在垂体和乳腺中高表达,在其他组织的表达特异性有待进一步研究.     

Enhancement of bovine growth hormone gene expression by increasing the plasmid copy number

Effect of copy number on the expression of bovine growth hormone gene (bGH) was investigated using the copy number mutants such as pKBJ10, pBJ( tet)10, pUBJ10-1, and pUBJ10 plasmids. The cells harboring plasmids below 84 copies/cell did not produced detectable levels of bGH. When the ColE1 replicon was replaced with the mutated ColE1 replicon originated from pUC19 plasmid, the copy number was increased to about 300 copies/cell and bGH production was enhanced by 11.5% (pUBJ10-1) and 12.3% of total cell protein (pUBJ10). A large amount of mRNA caused by increment of copy number would be needed to overcome some inhibitory threshold and might be an important factor for regulating bGH expression.     

牛呼吸道合胞体病毒G蛋白的截短表达与鉴定

经生物学软件DNA Star分析,将牛呼吸道合胞体病毒G基因截短成2个片段G1和G2。然后用人工合成的牛呼吸道合胞体病毒G基因为模板,用PCR分别扩增G1和G2基因片段,其大小分别为570 bp和308 bp。将目的片段定向克隆到pET30a表达载体中,酶切及测序鉴定均正确后,转化BL21表达菌,经IPTG诱导后G1和G2基因片段都获得了表达,且都为可溶性表达。用Ni柱亲和层析法在非变性条件下纯化重组蛋白,经免疫印迹试验鉴定证明纯化的重组蛋白G1具有良好的抗原性和特异性,而重组蛋白G2无反应性。应用纯化的重组蛋白G1进行的间接ELISA与免疫印迹试验在国内牛血清中检测到了BRSV血清抗体。本研究所表达的重组蛋白G1为基于牛呼吸道合胞体病毒G蛋白的血清学诊断方法的建立与牛呼吸道合胞体病毒G蛋白生物学功能的研究奠定了基础。     

Convergent occurrence of the developmental hourglass in plant and animal embryogenesis?

Background The remarkable similarity of animal embryos at particular stages of development led to the proposal of a developmental hourglass. In this model, early events in development are less conserved across species but lead to a highly conserved ‘phylotypic period’. Beyond this stage, the model suggests that development once again becomes less conserved, leading to the diversity of forms. Recent comparative studies of gene expression in animal groups have provided strong support for the hourglass model. How and why might such an hourglass pattern be generated? More importantly, how might early acting events in development evolve while still maintaining a later conserved stage?Scope The discovery that an hourglass pattern may also exist in the embryogenesis of plants provides comparative data that may help us explain this phenomenon. Whether the developmental hourglass occurs in plants, and what this means for our understanding of embryogenesis in plants and animals is discussed. Models by which conserved early-acting genes might change their functional role in the evolution of gene networks, how networks buffer these changes, and how that might constrain, or confer diversity, of the body plan are also discused.Conclusions Evidence of a morphological and molecular hourglass in plant and animal embryogenesis suggests convergent evolution. This convergence is likely due to developmental constraints imposed upon embryogenesis by the need to produce a viable embryo with an established body plan, controlled by the architecture of the underlying gene regulatory networks. As the body plan is largely laid down during the middle phases of embryo development in plants and animals, then it is perhaps not surprising this stage represents the narrow waist of the hourglass where the gene regulatory networks are the oldest and most robust and integrated, limiting species diversity and constraining morphological space.