This paper describes the ultrasound-guided transvaginal method of oocyte collection in cattle. Both technical and physiological aspects affecting the efficiency of the method are presented. The results of the author's experiments on oocytes collection and their developmental competence is also discussed.
Two procedures for mass production of embryos for use in cattle breeding schemes are now available, namely multiole ovulation + embryo transfer and more recently in vitro embryo production. Hormonal treatment for multiple ovulation, nonsurgical embryo collection and embryo transfer are widespread techniques to obtain more offspring from genetically superior cattle (MOET programme).However, the costs can be considerable and the yield of embryos is highly variable.Research during the past decade has been focused on embryo production in vitro with oocytes from slaughterhouse ovaries and has tremendously succeed.Presently ultrasound guided technique for oocyte collection in living animals has been developed.However, both discussed procedures allow to use only a tiny part of high follicular capital of ovary.Bovine ovary contains many thousands of primordial follicles, but the vast majority become atretic during growth and maturation.Recently is developed a technique for in vitro growth of prenatal follicles rescuing them against atresia.It would offer a significant means for the propagation of valuable animal stocks and would be an addition to the methods already available for use in embryo production in vitro, since it would supply a large and uniform population of oocytes from genetically superior animals.
Two cattle chromosome painting probes, identifying X and Y heterosomes, were applied to verify the diagnosis of XXY trisomy in an 8-month-old bull of the Polish Red breed. The probes were obtained after chromosome microdissection and labelled with biotin-16-dUTP. In all metaphase spreads, three fluorescence signals were observed - two X and one Y - confirming the diagnosis of a pure XXY trisomy.
Polymorphism of 11 microsatellite DNA loci was analysed in Polish Red (PR), Hereford and Holstein-Friesian (HF) cattle raised in Poland and genetic distance among these breeds was determined. At the 11 loci (TGLA227, BM2113, TGLA53, ETH10, SPS115, TGLA126, TGLA122, INRA23, ETH3, ETH225 and BM1824) analysed with automated DNA sizing technology, a total of 213 alleles were identified: 76 in PR, 76 in HF, and 61 in Hereford. All the microsatellite DNA markers showed high polymorphism. Polymorphism information content (PIC) calculated for each marker exceeded 0.5, except for the ETH3 locus in Hereford cattle (PIC=0.475), and heterozygosity (H) ranged from 54.1% to as much as 85.2%. The coefficient of genetic distance was 0.354 between PR and Hereford, 0.414 between HF and Hereford, and 0.416 between PR and HF cattle.
Attending animals of only one sex, depending on the chosen type of production, would considerably increase its effectivity. The problem of sex determination is an element of crucial importance in biotechniques of unconventional reproduction. In the case of cattle it gained importance with the development of breeding programmes based on multiple ovulation and embryo transfer (MOET). The control of proportions between sexes may be obtained either through separation (sorting) of spermatozoa subpopulations carrying chromosome X or Y, or by determining the sex of embryos (known as sexing) prior to their transfer to the uterus of the recipient cow. The current possibilities of sorting viable sperm are highly unsatisfactory for the needs of artifical insemination. Thus, for the sex preselection DNA techniques are being introduced, using the latest results on sex determining region Y (SRY) and on polymorphism of the gene of amelogenine. Introducing embryo-sexing into breeding programmes for cattle would result in considerable profits for the countries' economy. For instance, the annual genetic progress as regards milk production would reach 3 kg on the dam-son path and as much as 44 kg on the dam-daughter path.
Due to the functions that estrogens play in the regulation of reproduction, development of the mammary gland, growth and differentiation of cells, estrogen receptors and their genes are considered candidates for the markers of production and functional traits in farm animals, including cattle. In the present study, on the basis of the sequences of the human, ovine, and porcine ER genes, available in the GenBank database, sets of PCR primers were designed and used to amplify the bovine ERa gene 5?-region. Seven overlapping fragments of the 5? region of the bovine ERa gene were amplified and then sequenced. Altogether, these fragments were composed in the 2853-bp sequence which was deposited in the GenBank database under accession no. AY340597. The sequenced fragment included the noncoding exons A, B, C, their putative promoters, and a part of the coding exon 1. A polymorphism within the 5? region of the bovine ERa gene ? A/G transition, which could be recognized with RFLP-BglI, lying upstream to the exon C, was identified for the first time using this sequence.
Cattle breeding program for improvement of milk traits is accompanied by intensive changes in the structural and functional specificity of the animal organism. Assuming the hypothesis that the biological role of the female is to rear her progeny, it may be concluded that the extremely high milk productivity of the modern cow many-fold exceeds the physiological normal range. The mammary gland as a milk-producing highly effective bioreactor is exposed to the particularly strong influence of external and internal factors. Therefore, susceptibility to udder dysfunction generally called 'mastitis' causes great economical losses in highly productive cows. Mastitis is usually induced by a bacterial infection conveyed through the teat canal. The high variability of pathogens and diversity of environmental conditions cause difficulties in mastitis treatment. Antibiotic therapy does not give satisfactory results. Scientific research aims to recognize the heritable specificity of organism defence systems. Still, the currently used breeding selection procedures cannot be successful because natural resistance treated in categories of quantitative genetic variation shows a very low heritability and non-additive genotype-environment interaction. To overcome this problem, an alternative approach to detect a single gene with a high protective expression can be effective. The topics presented in this review include expression of lysozyme and lactoferrin in mammary gland tissue regarded as candidate gene for mastitis resistance as well as BoLA histocompatibility complex and milk protein polymorphic systems proposed as potential genetic markers of natural resistance in cattle.
Suboptimal conditions of oocyte maturation and embryo culture in vitro reduce their developmental competence as compared to in vivo development. It is well known now that some IVF blastocysts, despite a proper morphology, are not able to hatch and implant after transfer. Therefore, it is suggested that during in vitro culture embryos show an ability to adjust to suboptimal conditions, which is however accompanied by the changes in gene expression pattern resulting in reduced quality. Although gene expression analysis, as an invasive method, is considered as an indirect way of embryo quality evaluation, it provides valuable data on the processes crucial for growth and development.
Technology of bovine in vitro embryo production (IVP) had developed during the last few years to the level which allows its practical applications. Recently, IVP technology has been commercially used in several countries to increase the number of calves originating from donors of high genetic value. Combined with ovum pick-up technique (OPU), IVP technology serves as en alternative or integral method in breeding program MOET. The technology of IVP includes three developmental steps, i.e. oocyte in vitro maturation, in vitro fertilization and embryo culture. This review will focus on immature bovine oocyte recovery and selection, in vitro maturation, sperm processing, embryo culture to hatched blastocysts stage and factors affecting IVF efficiency. Moreover, recently developed investigations (IVP using calf oocytes and oocytes originating from small ovarian follicles) aiming to increase the effectiveness of IVP technology are discussed as well.
The main purpose of nuclear transfer in domestic species is to produce a large number of identical animals. There are two main ways to produce clones by nuclear transfer. One is to use the ICM and ED cells cultured under special conditions, as donor nuclei. The other way is to use the nuclear transfer embryo itself as the donor for the next generation of cloning (multiple generational cloning). The in vitro and in vivo developmental ability of nuclear transferred embryos is the same in the case of the first three generations. A limited number of multiple-generation clones were transferred into recipient heifers, resulting in offspring from I, II and III generation clones. The strategy to increase the efficiency of multiple generational bovine embryo cloning is discussed as well as the possible use of rabbit embryos as an experimental model.
Data included 393 097 calving ease, 129 520 gestation length, and 412 484 birth weight records on 412 484 Gelbvieh cattle. Additionally, pedigrees were available on 72 123 animals. Included in the models were effects of sex and age of dam, treated as fixed, as well as direct, maternal genetic and permanent environmental effects and effects of contemporary group (herd-year-season), treated as random. In all analyses, birth weight and gestation length were treated as continuous traits. Calving ease (CE) was treated either as a continuous trait in a mixed linear model (LM), or as a categorical trait in linear-threshold models (LTM). Solutions in TM obtained by empirical Bayes (TMEB) and Monte Carlo (TMMC) methodologies were compared with those by LM. Due to the computational cost, only 10 000 samples were obtained for TMMC. For calving ease, correlations between LM and TMEB were 0.86 and 0.78 for direct and maternal genetic effects, respectively. The same correlations but between TMEB and TMMC were 1.00 and 0.98, respectively. The correlations between LM and TMMC were 0.85 and 0.75, respectively. The correlations for the linear traits were above .97 between LM and TMEB but as low as 0.91 between LM and TMMC, suggesting insufficient convergence of TMMC. Computing time required was about 2 hrs, 5 hrs, and 6 days for LM, TMEB and TMMC, respectively, and memory requirements were 169, 171, and 445 megabytes, respectively. Bayesian implementation of threshold model is simple, can be extended to multiple categorical traits, and allows easy calculation of accuracies; however, computing time is prohibitively long for large models.
In the paper the detection of the SSCP polymorphism within the 5' fragment of bovine beta-lactoglobulin (LGB) gene is described. The 5' fragment of LGB gene (209 bp) was PCR-amplified and then subjected to electrophoresis allowing the detection of SSCP polymorphism. Among 124 animals (50 cows and 74 bulls) six SSCP patterns were identified and named R1, R2, R3, R4, R5 and R6, which occured with the frequency of 0.32, 0.51, 0.09, 0.06, 0.01and 0.01, respectively. The PCR-SSCP method is simple, fast, and relatively inexpensive. The SSCP polymorphism reported in the paper may be useful in looking for the associations between different SSCP patterns and LGB gene expression and milk properties.
Alloantibodies detecting antigenic determinats called BgC1 and AmiF1 were obtained in cattle.The analysis of determinants' inheritance was done on 1754 offsprings using 72 sires and 1740 dams.The molecule-carriers of both traits were characterized using serological and physico-chemical methods.It was found that identified determinants are markers of beta-globulin and alpha-microglobulin proteins, and of autosomal, dominant genes BgC1 and AmiF1 from independent loci which control them.
In this report we demonstrate a simple, effective and reliable diagnostic test of BLAD carrier detection based on specific PCR amplification of a 367 bp CD18 gene fragment and RFLP analysis using Taq I restriction enzyme. In a non random population of 220 animals we found 48 BLAD carriers. Within the amplified PCR fragment an unknown intron sequence of 159 bp was identified.
The bovine beta-lactoglobulin (LGB) gene is considered a potential quantitative trait locus in dairy cattle breeding. In Black-and-White dairy cattle the LGB gene has two predominant alleles A and B. This can result in three possible genotypes AA, AB and BB. Moreover, within the promoter of the gene several point mutations were found. A herd of one hundred and twenty-four Black-and-White cows were genotyped for two loci: locus LGB (exon IV, alleles A and B) and locus LGB-R (SSCP polymorphism within a fragment of LGB promoter: SSCP patterns R2, R3, R1, R9). In our sample 13 AA, 58 AB and 53 BB LGB cows and 66 R2, 16 R3, 40 R1 and 2 R9 LGB-R cows were identified. A statistical analysis revealed significant associations between LGB, LGB-R genotypes as well as intragenic haplotypes LGB/LGB-R and milk protein content during the first complete lactation. Cows with AA LGB genotype, R3 LGB-R SSCP pattern and AA/R3 haplotypes had the highest protein content. These results support the hypothesis that sequence variation within the promoter of the LGB gene is probably one of the factors responsible for differences in milk protein content.
Expression patterns of candidate genes with important functions in animal metabolism can help to identify potential molecular markers for cattle production traits. Reverse transcription followed by polymerase chain reaction is a method for rapid and accurate mRNA quantification. However, for exact comparison of mRNA quantity in various samples or tissues, it is important to choose appropriate reference genes. In cattle, little information is available on the expression stability of housekeeping genes (HKGs). The aim of the present study is to develop a set of reference genes that can be used for normalization of concentrations of mRNAs of genes expressed in the bovine liver, kidney, pituitary and thyroid. The study was performed on 6-, 9-, and 12-month-old bulls of dairy and meat cattle breeds. Six HKGs were investigated: ACTB, GAPDH, HPRTI, SDHA, TBP, and YWHAZ. The most stably expressed potential reference HKGs differed among tissues/organs examined: ACTB, TBP, YWHAZ, GAPDH, HPRTI, and SDHA in the liver; GAPDH and YWHAZ in the kidney; GAPDH and SDHA in the pituitary; and TBP and HPRTI in the thyroid. The results showed that the use of a single gene for normalization may lead to relatively large errors, so it is important to use multiple control genes based on a survey of potential reference genes applied to representative samples from specific experimental conditions.
Doublesex and mab-3 related transcription factor 1 (DMRT1) is considered to be the most conserved gene among loci involved in the molecular pathways of animal sexual development. In the majority of the extensively examined vertebrates, its function is limited to the upstream or downstream testis regulators acting during embryogenesis. Our present study demonstrated the structural homology between DMRT1 orthologos in human and cattle. A BAC clone with a specific bovine sequence of the gene was used in the FISH mapping experiments. The physical localization of DMRT1 in cattle (BTA 8q17) was determined and its homology to the human locus was shown (HSA 9p24.3). Furthermore, another BAC probe, containing the sequence of the human homologue (pBACe3.6), generated hybridisation signals on bovine metaphase chromosomes and indicated the physical location of the autosomal bovine DMRT1 locus. Further investigations of the gene in domestic animals might provide more support for its conservative status and may help in understanding the molecular mechanisms involved in the occurrence of sexual abnormalities often diagnosed in livestock.
Fluorescence in situ hybridization (FISH) experiments with specific probes for chromosome 29 and 25 were carried out on a Brown Swiss bull, previously diagnosed as a carrier of 1;29 centric fusion.The hybridization of the chromosome 29-specific probe (BMC 4216-already located on 29q13), produced signals on two small acrocentrics, but not the translocated chromosome.The signals appeared on the translocated chromosome and on a single chromosome 25 after hybridization of the chromosome-specific probe (BMC 3224 -previously located on 25q24).According to the actual nomenclature, the analysed aberration is a robertsonian translocation involving chromosomes 1 and 25.
Twinning in cattle ranges from about 1% for beef breeds to about 4% for dairy breeds. The incidence of double births may have both positive and negative effects, which mainly depends on the purpose for which the cattle are raised. Because of freemartinism, as well as management problems connected e.g. with a greater risk of dystocia and retained placenta, it is an undesirable trait in dairy herds. In beef cattle, however, twinning can considerably increase the efficiency of production. Low heritability, a long generation interval for progeny testing, sex-limited expression and an unfavourable correlation with milk yield make twinning difficult to control by selection. Hence, it is the type of trait for which the identification of the genetic marker - quantitative trait loci (QTL) linkage and the implementation of marker-assisted selection in breeding strategies are expected to be especially beneficial. Searching for QTL influencing the reproductive rate in cattle was performed mainly in the US Meat Animal Research Center twinning herd and in the commercial Norwegian cattle population. Among several genome regions that appear to control twinning and ovulation rates, the most interesting seem to be chromosomes 5, 7, 19 and 23.
We report on a PCR-RFLP procedure for recognising of a silent point mutation of ITGB2 CD18 subunit gene in cattle. Polymorphism screening was performed in a Polish Black-and-White cattle population (n=210). The genotype and allele frequecies were established in the sires and cows. Further research is needed to explain the possible applications of the CD18 silent point mutation as a potential molecular marker for high milk productivity.
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