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20192
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Subject: Animal Biochemistry × End date: 2019 × Start date: 2019 ×
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    ThesisItemOpen Access
    CHARACTERISATION OF GENES AFFECTING METABOLIC ADAPTATION IN TRANSITION DAIRY COWS
    (College of Veterinary and animal Science,Mannuthy, 2019) VASUDHAR BHAT S.V.; Shynu M.
    The period from three weeks before parturition until three weeks after parturition is the transition period in dairy cows. This time period is critical in the health and production of dairy cows and profitability of dairying as most diseases occur during this period. The dairy cows undergo several metabolic adaptations to overcome the stress of foetal growth and lactation. Even when maintained under similar feeding and managemental conditions, dairy cows exhibit variation in adaptability. This indicates the genetic influence on metabolic adaptability. The present study was performed in 30 apparently healthy pregnant crossbred dairy cows with an objective to characterize three genes responsible for metabolic adaptability and to detect variations in these genes, among animals with varying adaptability, as indicated by their BHBA concentration. Blood was collected and serum was separated at fortnightly intervals from four weeks before the predicted day of parturition until four weeks after parturition. Concentration of NEFA and BHBA were determined. The mean concentration of NEFA (0.254±0.21 mmol/L) and BHBA (0.608±0.024 mmol/L) during pregnancy were significantly elevated in comparison with the NEFA (0.349±0.37 mmol/L) and BHBA (0.681±0.23 mmol/L) during lactation. As BHBA is a better indicator of metabolic adaptability, the animals were grouped based on their BHBA concentration using cluster analysis. The genes DNAJC30, SNAI2 and UEVLD known to influence metabolic adaptability were sequenced from representative animals of each group. Four differences (transitions) in sequence were observed downstream the coding region of DNAJC30 gene between the groups. One change was located in the promoter region and another in exon one of SNAI2 gene. The CDS of UEVLD gene did not show any change when compared with the available Bos taurus sequence in NCBI. The variations in genes studied might contribute to difference in the metabolic adaptability between the groups. Further studies in a larger population, is required to ascertain the suitability of using these as markers for selection of dairy animals for better metabolic adaptability
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    ThesisItemOpen Access
    ASSESSMENT OF IMMUNOMODULATORY EFFECTS OF GOAT LACTOFERRIN AND ITS PEPSIN HYDROLYSATE
    (College of Veterinary and animal Science,Mannuthy, 2019) DEEPAK CHANDRAN; UMA R.
    Lactoferrin, well known as a minor whey protein, is an 80kDa iron-binding glycoprotein primarily present in milk. It exhibits an array of biological activities including antioxidant, antimicrobial, anticancer, metal binding and immunomodulatory properties. Many studies point out that whey proteins contain various bioactive peptides in their primary sequences whose immunoregulatory and therapeutic potentials have not been much explored. The present study focussed on the isolation and characterisation of lactoferrin (gLf) from the colostrum/milk of Malabari goats, followed by the preparation of its pepsin hydrolysate (gLPH) so as to assess their in vitro immunomodulatory potentials on peripheral blood mononuclear cells (PBMCs). Colostrum samples collected from Malabari goats maintained at University Goat and Sheep Farm, College of Veterinary and Animal Sciences, Mannuthy were processed and treated with ammonium sulphate to remove globulins from the samples. Fractions containing albumin and remaining proteins including lactoferrin after dialysis were loaded on to CM- Sephadex C-50 cation exchanger column and eluted with a step gradient of 0.4, 0.6 and 0.8M NaCl. The presence of gLf in the high OD280 value fractions eluted with 0.8M NaCl was confirmed by 12 per cent SDS-PAGE and Western blotting. The concentration of gLf as estimated by Lowry’s method was found to be 15.103 mg/L of colostrum. The gLf was hydrolysed by treatment with three per cent porcine pepsin under acidic conditions at 37 °C for four hours to form gLPH. The immunomodulatory potentials of gLf and gLPH were studied on bovine PBMCs which were cultured along with specific combinations of mitogen, phytohaemagglutinin (PHA)/gLf/gLPH at 37°C for 72h in five per cent CO2 humidified air. A wide range of concentrations of both gLf and gLPH were utilized to assess their cytoproliferative effect on PBMCs with or without mitogens. In the presence and absence of mitogen, higher concentrations of lactoferrin were found to significantly inhibit the proliferation of PBMCs whereas lower concentrations brought about significantly active proliferation of the cells. Maximum proliferation with or without mitogen was observed with 1.5 µg/mL of culture. The cell proliferation potentials of gLPH was higher than that of gLf. With gLPH, in the presence and absence of mitogen, maximum proliferation of PBMCs could be detected with the highest concentration of 50 µg/mL. The cells cultured with gLf and gLPH showing maximum cell proliferation were harvested and their RNA were isolated to study the expression of pro-inflammatory cytokines by real time PCR. It was demonstrated that gLf and gLPH showed potentiated anti-inflammatory activity by significantly inhibiting the expression of the pro-inflammatory cytokine genes IL1β, IL-6 and TNF-α.