Analiza učestalosti goveđeg beta-kazeina tipa A1 i A2 u krava holštajnsko-frizijske pasmine PCR-om u stvarnom vremenu s fluorescentnim hibridizacijskim sondama

A2 milk popularity is increasing across the world and novel molecular techniques have been evaluated to develop reliable methods. This study aimed to genotype Holstein-Friesian cows concerning their A1/A2 status using Real-time PCR assay with the specifically designed FRET hybridization probes. In this context, DNA samples were obtained from 310 Holstein-Friesian milk samples. Concerning the Real-time PCR assay, the melting temperature of each amplicon was analyzed and the melting data was converted to a derivative plot using the LightCycler 480 System. The sensor probe was designed to match the wild-type sequence in the target DNA. In the Real-time PCR assay, the melting peaks obtained in the Real-time PCR assay were highly decisive and consistent for each genotype regarding CCT→CAT alteration. The results indicated a remarkably high frequency of the A2 allele (68%) and a considerable frequency of heterozygous animals (0.41). Population genetic analysis showed intermediate levels of genetic variability and biodiversity. The A2-herd conversion process is a complex process consisting of genetic testing of both cows and calves, evaluating replacement rates, and the conversion of heterozygotes by using A2-genotyped bull semen. In this sense, the key point is a reliable and rapid genotyping method to produce A1-free milk. This study suggests that Real-time PCR assay with the specifically designed FRET hybridization probes is a preferable method for A2 genotyping, and may be useful for further studies and instructive for companies or breeders who aim to produce A2 milk.Popularnost mlijeka s A2 tipom beta-kazeina sve je veća diljem svijeta što je popraćeno razvojem novih i pouzdanijih metoda za molekularne analize. Cilj je ovog istraživanja bila genotipizacija holštajnsko-frizijskih krava s obzirom na A1/A2 status primjenom PCR-a u stvarnom vremenu (RT-PCR) s posebno dizajniranim FRET hibridizacijskim sondama. DNA je dobivena iz uzoraka mlijeka 310 krava. RT-PCR-om analizirana je temperatura taljenja svakog amplikona te su ti podaci uz upotrebu sustava LightCycler 480 korišteni za iscrtavanje grafičkih prikaza. Senzorska sonda dizajnirana je tako da odgovara divljem tipu sekvencije u ciljnoj DNA. Vršne vrijednosti taljenja dobivene RTPCR-om bile su postojane razlučivosti za svaki genotip s obzirom na promjenu CCT→CAT. Rezultati su pokazali vrlo visoku učestalost alela A2 (68 %) i znakovitu učestalost heterozogotnih životinja (0,41). Genetska analiza u populaciji pokazala je srednje vrijednosti genetske varijabilnosti i bioraznolikosti. Proces usmjeravanja uzgoja prema životinjama s A2 tipom mlijeka složen je proces koji uključuje genetsko testiranje krava i teladi, procjenu stopa zamjene i konverziju heterozigota upotrebom A2-genotipiziranog sjemena bikova. U tom je smislu ključna brza i pouzdana metoda genotipizacije životinja koje proizvode mlijeko bez A1 tipa beta-kazeina. Ovo istraživanje pokazuje da se RT-PCR s posebno dizajniranim hibridizacijskim FRET sondama može preporučiti za A2 genotipizaciju goveda. Navedena metoda mogla bi biti korisna u daljnjim istraživanjima te u industriji i u uzgoju stada za proizvodnju mlijeka s A2 tipom beta-kazeina

New palladium(II) and platinum(II) 5,5-diethylbarbiturate complexes with 2-phenylpyridine, 2,2’-bipyridine and 2,2’-dipyridylamine : synthesis, structures, DNA binding, molecular docking, cellular uptake, antioxidant activity and cytotoxicity

Peer reviewedPostprin

Procjena nekih čimbenika okoliša i genetskih čimbenika (CSN3 i AGPAT6) na prinos i sastav mlijeka u sanske koze

This study was designed to determine the effect of some environmental factors on milk yield and composition in Saanen goats and the effect of CSN3 and AGPAT6 gene polymorphisms on milk production traits. Saanen goats were genotyped for milk traits, and the samples were collected during the lactation, and the milk yield/compositions of each goat were specified. In terms of the CSN3 and AGPAT6 genes, the genotypes were defined by PCR-RFLP. The milk yield (MY), lactation length (LL), protein, fat, total solid (TS), solid not fat (SNF), casein, lactose rates and yields were as follows; 388.9 ± 17.5 kg, 243 ± 5.81 days, 3.29 ± 0.05%, 3.59 ± 0.07%, 11.60 ± 0.13%, 8.43 ± 0.06%, 2.57 ± 0.04%, 4.31 ± 0.03%, 11.21 ± 0.47 kg, 12.79 ± 0.64 kg, 40.95 ± 1.82 kg, 29.53 ± 1.31 kg, 8.82 ± 0.37 kg, 15.40 ± 0.74 kg respectively. The determined genotype and allele frequencies of CSN3 and AGPAT6 gene were as follows; FF (%99.20), MF (%0.80), F (0.996), M (0.004); GG (%12), GC (%43.20), CC (%44.80), G (0.336), C (0.664). The impacts of lactation length on all milk yield parameters (protein, fat, TS, SNF, casein, lactose, MY) (PG), the herd was found to be monomorphic (FF) for the CSN3 gene. The effect of the AGPAT6 gene on milk traits in Saanen goats was determined not to be statistically significant. Environmental factors, such as maternal age and lactation length, were found to significantly affect some milk traits in Saanen goats. So consideration of factors such as maternal age or lactation length could be useful for improving breeding strategies for dairy goats.Istraživanje je provedeno kako bi se odredili učinci čimbenika okoliša te učinci genskih polimorfizama CSN3 i AGPAT6 na prinos i sastav mlijeka u sanske koze. Tijekom laktacije koza provedena je genotipizacija te prikupljani uzorci za utvrđivanje prinosa i sastava mlijeka. Genotipovi gena CSN3 i AGPAT6 određeni su PCR-RFLP-om. Prinos mlijeka (MY), duljina laktacije (LL), udio proteina, udio masnoće, udio suhe tvari (TS), udio nemasne suhe tvari (SNF), udio kazeina, udio laktoze bili su kako slijedi: 388,9 ± 17,5 kg, 243 ± 5,81 dana, 3,29 ± 0,05 %, 3,59 ± 0,07 %, 11,60 ± 0,13 %, 8,43 ± 0,06 %, 2,57 ± 0,04 %, 4,31 ± 0,03 %. Prinos u kg za protein, masnoću, suhu tvar, nemasnu suhu tvar, kazein i laktozu bili su kako slijedi: 11,21 ± 0,47 kg, 12,79 ± 0,64 kg, 40,95 ± 1,82 kg, 29,53 ± 1,31 kg, 8,82 ± 0,37 kg, 15,40 ± 0,74 kg. Utvrđeni genotipovi i učestalost alela gena CSN3 i AGPAT6 bili su: FF (99,20 %), MF (0,80 %), F (0,996), M (0,004), GG (12 %), GC (43,20 %), CC (44,80 %), G (0,336), C (0,664). Statistički znakoviti utjecaji utvrđeni su za: duljinu laktacije na pokazatelje prinosa mlijeka (proteine, masnoću, TS, SNF, kazein, laktozu, MY) (P G), dok je stado za gen CSN3 bilo monomorfno (FF). Učinak gena AGPAT6 na svojstva mlijeka nije bio statistički znakovit. čimbenici kao što su dob majke i duljina laktacije znakovito su utjecali na svojstva mlijeka sanske koze, stoga bi oni mogli biti korisni za unapređenje uzgojnih strategija mliječnih pasmina koza

Effect of STAT1, OLR1, CSN1S1, CSN1S2 , and DGAT1 genes on milk yield and composition traits of Holstein breed

ABSTRACT The objectives of the study were to evaluate allelic frequencies and test the association between STAT1, OLR1, CSN1S1, CSN1S2 , and DGAT1 gene polymorphisms and milk production traits including lactation milk yield, 305 days milk yield, days before peak milk production, and peak milk yield. In addition, samples of milk were analysed for fat, protein, lactose, and total solid contents. A total of 168 purebred Holstein-Fresian cows were genotyped using polymerase chain reaction and restriction fragment length polymorphism methods. Statistical analysis was carried out using least square methods of the general linear model procedure. Significant differences were found between genotypes of the CSN1S2 locus in relation to milk protein content. In addition, DGAT1 was significantly associated with peak milk production. These results suggested that CSN1S2 and DGAT1 markers may be evaluated to achieve various commercial goals in dairy cattle production

Low-penetrance susceptibility variants and postmenopausal oestrogen receptor positive breast cancer

WOS: 000513636700001The risk of breast cancer (BC) in women is high and many factors including genetic factors increase the risk for the disease. It is revealed that the variations of low-penetrance susceptibility genes are important for carcinogenesis as they interact with the environmental and hereditary factors. Recently, the list of BC-associated common single nucleotide polymorphisms (SNPs) and chromosomal loci in low-penetrance susceptibility genes have been expanded in genomewide association studies. FGFR2, LSP1, MAP3K1, TGFB1, TOX3, 2q35 and 8q loci variations are some examples for these common SNPs. These SNPs and their association with BC risk was investigated in many different populations. Therefore in this study, we aimed to evaluate low-penetrance susceptibility SNPs; namely FGFR2 rs1219648, rs2981579, rs2981582; MAP3K1 rs889312; TOX3 rs3803662; LSP1 rs909116, rs3817198 and SLC4A7 rs4973768 together, for the first time in Turkish postmenopausal oestrogen receptor positive BC cases. Following the DNA isolation, multiplex PCR and matrix-assisted laser desorption/ionization mass spectrometry with time of flight measurement (MALDI-TOF) based SNP analysis were performed. MAP3K1 rs889312 SNP demonstrated the strongest association with BC risk among the other low penetrant SNPs, it was also associated with BC risk in a dominant model. Only in a ressesive model, TOX3 rs3803662 was associated with BC risk. In addition, rs4973768 CC and rs909116 CC genotypes are correlated with higher tumour size which is not reported in the literature as yet; on the other hand there are no associations between any of the other SNP genotypes and clinopathological parameters. In our opinion, MAP3K1 rs889312 may be a good BC susceptibility biomarker candidate for Turkish population.Kastamonu University Scientific Research Projects Management Co-ordination OfficeKastamonu University [KuBAP-01/2013-02]This study was supported by the Kastamonu University Scientific Research Projects Management Co-ordination Office with code Number KuBAP-01/2013-02

An investigation of the effects of FGFR2 and B7-H4 polymorphisms in breast cancer

Introduction: Polymorphisms in FGFR2 are important markers for breast cancer susceptibility in the general population. CHEK2 and FGFR2 polymorphisms with known susceptibility alleles of BRCA1, BRCA2, PTEN, and TP53, can be investigated as potential modifiers of high penetrant risk alleles. Although the B7-H4 gene is highly expressed in many different tumors, there is one published study showing the association of polymorphisms with breast cancer. We aimed to investigate FGFR2 and B7-H4 polymorphisms in breast cancer in the Turkish community. Materials and Methods: In a group of 31 cases diagnosed with breast cancer and 30 healthy women with matched ages, the single-nucleotide polymorphisms (SNPs) rs1219648, rs2981582 in FGFR2 gene were identified by sequence analysis and the SNPs rs10754339, rs10801935, and rs3738414 in the B7-H4 gene were identified by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Statistical analysis was performed using SPSS. Results: Although statistically not significant, the frequency of FGFR2 heterozygous polymorphisms in the group with breast cancer was detected to be higher. In the B7-H4 SNP rs10801935, polymorphic AA, and AG genotype distributions were found in higher frequencies in the breast cancer patients. In contrast to the results of a published study, the present study shows that B7-H4 rs3738414 polymorphism GG genotype was found in higher frequency in the control group than the breast cancer group and the result was statistically significant (P=0.018). Conclusion: Larger scale studies are necessary to determine the prevalence of these polymorphisms and association with breast cancer in Turkish community, as this study is the first study performed

Increased Serum Osteoprotegerin Levels Associated with Decreased Bone Mineral Density in Familial Mediterranean Fever

Familial Mediterranean fever (FMF) has episodic or subclinical inflammation that may lead to a decrease in bone mineral density (BMD). The aim of this study was to evaluate the effect of FMF on bone metabolism and to investigate the factors that can influence bone metabolism, such as body mass index (BMI), mutations in Mediterranean fever (MEFV) gene, osteoprotegerin (OPG), leptin and inflammatory cytokines, including interleukin (IL)-1 beta, IL-6 and tumor necrosis factor-alpha (TNF-alpha). OPG, a soluble protein produced by osteoblasts, favors increased bone mass. Leptin may influence bone metabolism by acting on differentiated osteoblasts, having anabolic effects on bone. Thirty-one FMF patients in attack-free period (12 females and 19 males; mean age 31.4 +/- 9.3 years) and 18 healthy controls (11 females and 7 males; mean age 34.6 +/- 9.5 years) were compared according to the above parameters. BMD (g/cm(2)) and standard deviation scores (Z-score) were measured at the lumbar spine L-1-L-4 (BMD-L1-4) and proximal femur by dual X-ray absorptiometry. Osteopenia is defined as a Z-score between -1 and -2.5 and osteoporosis is equal or below -2.5. FMF patients showed statistically significant reduction in BMD-L1-4 and Z-score-L1-4. Moreover, serum OPG concentration was significantly elevated in FMF patiens. In contrast, MEFV gene mutations, leptin and the inflammatory cytokines did not differ between the patient and control groups. In conclusion, BMD was decreased and OPG was increased in our FMF patients. The high OPG levels may reflect a preventive mechanism against bone loss; namely, OPG might protect the FMF patients from excessive osteoporosis

Association of polymorphisms in lipid and energy metabolism-related genes with fattening performance in Simmental cattle

Lipid and energy metabolism are major constituents of mammal growth and thus fattening performance of cattle. This study was designed to evaluate the effects of polymorphisms in lipid and energy metabolism-related genes including oxidized low-density lipoprotein receptor 1 (OLR1), lactoferrin (LTF), stearoyl-CoA desaturase (SCD), beta-lactoglobulin (LGB), thyroglobulin (TG), annexin A9 (ANXA9), myogenic factor 5 (MYF5), protein kinase AMP-activated non-catalytic subunit gamma 3 (PRKAG3), and pituitary-specific transcriptional factor 1 (PIT1), on fattening performance in Simmental cattle. A total of 72 purebred Simmental bulls with a similar initial age and weight were fattened on the same farm for 10 months. Association analysis was performed using linear mixed models. The OLR1 marker was significantly associated with the final weight (FW), hot carcass weight (HCW), chilled carcass weight (CCW), dressing percentage (DP), and total weight gain (TWG). SCD affected the FW, TWG, and average daily live weight gain (ADWG). The present results clearly demonstrated the significant impact of the TG marker on fattening performance. It was highly significantly associated with the FW, HCW, CCW, and TWG. The SCD × TG and the OLR1 × TG interactions had remarkable effects on the traits analyzed. The GACC and CCCC haplotypes of the SCD × TG and OLR1 × TG, respectively, were found to be powerful markers for fattening performance in Simmentals. Novel associations in this study may be useful for further genetic evaluations to improve beef cattle breeding.</p