Polymorphism of α_S1-casein in goat milk: identification of A, B, E and F variants by biochemical and genetic analysis

Many researches, related to genetic polymorphism of αS1-casein in goat milk, showed a marked variability and implications in milk and dairy product traits. The genetic variants (from A to G) were associated with four levels of expression: 3,6 g/L per A, B and C (“strong” alleles); 1,6 g/L per E (“medium” allele); 0,6 g/L per F (“weak” allele) and 0 g/L per O (“null” allele). Differences existing among the A, B, C, D, E, F, G and O have been described (Grosclaude et al., 1994). To analyze genetic polymorphism of caseins from goat milk, distinct electrophoretic (Russo et al., 1986; Addeo et al., 1988) and chromatographic techniques (Jaubert and Martin, 1992; Iametti et al., 1999) were applied. In last years, the advancement in technological field is allowing the research of primary structure of protein variants (Ferranti et al., 1997; Trujillo et al., 2000) and basic sequence of new alleles (Martin et al., 1999; Bevilacqua et al., 2002; Ramunno et al., 2002). In this work, we employed electrophoretic protein separation (IEF and SDS-PAGE), chromatographic analysis (RP-HPLC) and molecular biology techniques based on polymerase chain reaction (real-time PCR) to detect αS1-CN genetic variants from goat milk. The utility of protein and DNA analysis combination was discussed

Reciprocal Translocations in Cattle: frequency estimation

Chromosomal anomalies, like Robertsonian and reciprocal translocations represent a big problem in cattle breeding as their presence induces, in the carrier subjects, a well documented fertility reduction. In cattle reciprocal translocations (RCPs, a chromosome abnormality caused by an exchange of material between nonhomologous chromosomes) are considered rare as to date only 19 reciprocal translocations have been described. In cattle it is common knowledge that the Robertsonian translocations represent the most common cytogenetic anomalies, and this is probably due to the existence of the endemic 1;29 Robertsonian translocation. However, these considerations are based on data obtained using techniques that are unable to identify all reciprocal translocations and thus their frequency is clearly underestimated. The purpose of this work is to provide a first realistic estimate of the impact of RCPs in the cattle population studied, trying to eliminate the factors which have caused an underestimation of their frequency so far. We performed this work using a mathematical as well as a simulation approach and, as biological data, we considered the cytogenetic results obtained in the last 15 years. The results obtained show that only 16% of reciprocal translocations can be detected using simple Giemsa techniques and consequently they could be present in no less than 0,14% of cattle subjects, a frequency five times higher than that shown by de novo Robertsonian translocations. This data is useful to open a debate about the need to introduce a more efficient method to identify RCP in cattle

Polimorfizam κ-kazeina talijanskih pasmina koza: novi ACRS-PCR test za razlikovanje A i B alela

The objective of this study was to develop a DNA test for rapid characterisation of goat κ-casein (κ-CN) A and B variants and to study this polymorphism in Italian goat breeds. Genetic polymorphism of κ-CN gene was, in addition to isoelectric focusing, analysed according to a new technique designated as amplification created restriction site. Two alleles that differ in one nucleotide mutation (G Ø A) in exon 4 were characterised. The 167-bp PCR product surrounding the nucleotide mutation was amplified from genomic DNA and the PCR product was digested with MaeIII. After digestion the A allele gives three fragments of 77, 65 and 25 bp in comparison with the B allele which gives two fragments of 90 and 77 bp. The analysis of allele frequency distribution at κ-CN locus, based on 401 individual samples, revealed significant differences among three goat breeds from the north of Italy (Nera di Verzasca, Frontalasca and Alpine) with frequency of κ-CN B allele around 0.3, versus two goat breeds from the south of Italy (Maltese and Sarda) with frequency of κ-CN B allele around 0.5. While two goat breeds (Maltese and Nera di Verzasca) did not show significant deviations from the Hardy-Weinberg equilibrium, a highly significant excess of heterozygote genotype (AB) was observed in Alpine, Frontalasca and Sarda goats. Here the developed DNA method and observed relatively high frequency of κ-CN B allele give a prerequisite for the assessment of research related to the simultaneous estimation of the effects of composite αs1/κ-CN genotypes on milk production and cheese-making properties.Svrha je bila razviti DNA test za brzo određivanje genetičkih varijanti κ-kazeina u talijanskih pasmina koza. Genetički je polimorfizam istraživan izoelektričnim fokusiranjem (IEF) i amplifikacijom nastalim restrikcijskim mjestom na genomu (ACRS). PCR produkt na ekzonu 4, dug 167-bp koji okružuje nukleotidnu mutaciju (G Ø A), amplificiran s genomske DNA i razgrađen je s MaeIII enzimom. Identificirana su dva alela koja se razlikuju u jednoj nukleotidnoj mutaciji (G Ø A). Dobiveni su fragmenti 77 i 65 bp za alel A te 90 i 77 bp za alel B. Signifikantna razlika za frekvenciju alela B ustanovljena je za tri pasmine koza sa sjevera Italije (Nera di Verzasca, Frontalasca i Alpine) s frekvencijom κ-kazeina B oko 0,3, u usporedbi s dvjema pasminama s juga Italije (Maltese i Sarda), s frekvencijom κ-kazeina B otprilike 0,5. Pasmine Maltese i Nera di Verzasca nisu pokazale signifikantno odstupanje od Hardy-Weinbergove ravnoteže, za razliku od pasmina Alpine, Frontalasca i Sarda u kojima je uočeno veliko signifikantno odstupanje. Prikazana DNA metoda i promatrana relativno velika frekvencija κ-kazeina B alela omogućavaju daljnja istraživanja zajedničkog utjecaja αs1-kazeina i κ-kazeina na proizvodnju mlijeka i sira

Characterization of telomere length in Agerolese cattle breed, correlating blood and milk samples

Studies into telomere length in cattle are relatively recent and have focused mainly on the Holstein Friesian cattle breed, making it arduous to evaluate the correlation with ageing due to the early age of culling in this breed. Telomere length provides information about the productive lifespan and the quality of farm management, complying with the 'One Health' approach. This study evaluated telomere length in Agerolese cattle, an autochthonous dairy breed characterized by a long productive lifespan (13 years). Multiplex quantitative PCR estimated telomere length in DNA extracted from blood and milk matrices. Interestingly, the results showed longer telomeres in Agerolese (compared to the Holstein Friesian cattle control group), with a negative correlation between telomere length and increasing age and a synchronous trend between blood and milk samples, with a positive correlation between them

Successful treatment with T depleted autologous peripheral blood stem cell transplantation of refractory chronic autoimmune thrombocytopenic purpura

Autoimmune thrombocytopenia (AITP) is a disorder due to specific platelet auto-antibodies directed against platelet surface glycoproteins. AITP in adults is usually chronic, idiopathic and frequently refractory to conventional treatments. Myelo- and immuno- suppressive chemotherapy followed by autologous peripheral blood stem cell (PBSC) transplantation is an experimental approach for severe chronic refractory AITP. We report a case of a woman with AITP, refractory to the conventional therapy, submitted to T-cell-depleted autologous PBSC transplantation, which obtained long term stable response on platelet count. We deem that the positive outcome of our patient depends on T-cells depletion of the graft, which reduces autoreactive T clones

Clinical, genetic, and pathological features of male pseudohermaphroditism in dog

Male pseudohermaphroditism is a sex differentiation disorder in which the gonads are testes and the genital ducts are incompletely masculinized. An 8 years old dog with normal male karyotype was referred for examination of external genitalia abnormalities. Adjacent to the vulva subcutaneous undescended testes were observed. The histology of the gonads revealed a Leydig and Sertoli cell neoplasia. The contemporaneous presence of testicular tissue, vulva, male karyotype were compatible with a male pseudohermaphrodite (MPH) condition

Caprine αs1-Casein Polymorphism: Characterisation of A, B, E and F Variants by Means of Various Biochemical and Molecular Techniques

Considering a wide interest for the characterisation of caprine αs1-casein variants and a large number of differently equipped laboratories, the objective of this study was to analyse and compare characteristics of caprine αs1-casein variants by means of various biochemical and molecular techniques. The most frequent caprine αs1-casein variants (A, B, E and F) were characterized by employing electrophoretic protein separation analyses (capillary electrophoresis, isoelectric focusing, and sodium dodecylsulfate polyacrylamide gel electrophoresis), chromatographic analysis (reversed phase-high performance liquid chromatography) as well as DNA analyses (ASA and real-time polymerase chain reaction approach). Further, we stressed weak and strong points for each method applied and provided information for the optimal and complementary use of those methods with respect to time, resolution and costs

R-spondin1 and FOXL2 act into two distinct cellular types during goat ovarian differentiation

<p>Abstract</p> <p>Background</p> <p>Up to now, two loci have been involved in XX sex-reversal in mammals following loss-of-function mutations, PIS (Polled Intersex Syndrome) in goats and <it>R-spondin1 </it>(<it>RSPO1</it>) in humans. Here, we analyze the possible interaction between these two factors during goat gonad development. Furthermore, since functional redundancy between different <it>R-spondins </it>may influence gonad development, we also studied the expression patterns of <it>RSPO2, 3 </it>and <it>4</it>.</p> <p>Results</p> <p>Similarly to the mouse, <it>RSPO1 </it>shows a sex-dimorphic expression pattern during goat gonad development with higher levels in the ovaries. Interestingly, the PIS mutation does not seem to influence its level of expression. Moreover, using an RSPO1 specific antibody, the RSPO1 protein was localized in the cortical area of early differentiating ovaries (36 and 40 d<it>pc</it>). This cortical area contains the majority of germ cell that are surrounded by FOXL2 negative somatic cells. At latter stages (50 and 60 d<it>pc</it>) RSPO1 protein remains specifically localized on the germ cell membranes. Interestingly, a time-specific relocation of RSPO1 on the germ cell membrane was noticed, moving from a uniform distribution at 40 d<it>pc </it>to a punctuated staining before and during meiosis (50 and 60 d<it>pc </it>respectively). Interestingly, also <it>RSPO2 </it>and <it>RSPO4 </it>show a sex-dimorphic expression pattern with higher levels in the ovaries. Although <it>RSPO4 </it>was found to be faintly and belatedly expressed, the expression of <it>RSPO2 </it>increases at the crucial 36 d<it>pc </it>stage, as does that of <it>FOXL2</it>. Importantly, <it>RSPO2 </it>expression appears dramatically decreased in XX PIS^-/-gonads at all three tested stages (36, 40 and 50 d<it>pc</it>).</p> <p>Conclusion</p> <p>During goat ovarian development, the pattern of expression of <it>RSPO1 </it>is in agreement with its possible anti-testis function but is not influenced by the PIS mutation. Moreover, our data suggest that RSPO1 may be associated with germ cell development and meiosis. Interestingly, another RSPO gene, RSPO2 shows a sex-dimorphic pattern of expression that is dramatically influenced by the PIS mutation.</p