Identification of Polymorphisms of Gene CSN2 of B Casein in Greek Cow Breeds (Holstein) by Restriction Fragment Length Polymorphism

Vougiouklaki, Despina; Antonopoulos, Dionysios A.; Allexeli, Stella; Houhoula, Dimitra

doi:10.5539/jas.v12n11p32

Cited by 4 publications

(3 citation statements)

References 21 publications

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“…With respect to DNA extraction methods, methods 1 and 2 were a direct applications of a commercially available kits, NucleoSpin Tissue (Usman et al, 2014;Vougiouklaki et al, 2020) and NucleoSpin Blood (Macherey-Nagel, Düren, Germany), both kits are based on the binding of DNA to a silica gel membrane to get high-quality DNA. The main difference between them was the incubation time after adding proteinase K, which is was not necessary for NucleoSpin Blood and it was 2 h for the NucleoSpin Tissue kit.…”

Section: Dna Isolationmentioning

confidence: 99%

“…Apart from qPCR methods, in Mayer et al (2021) milk proteins were directly phenotyped through isoelectric focusing to discriminate A1 and A2 β-casein protein variants. However, most studies were focused on the genotyping of the CSN2 gene to determine the frequency of these genotypes in different cattle breeds (Masoumeh Firouzamandi et al, 2018;Mayer et al, 2021;Miluchová et al, 2014;Rangel et al, 2017;Vougiouklaki et al, 2020). Identification of animal genotypes may not be enough to A2 milk authentication (Giglioti et al, 2020).…”

Section: Sensitivity: Limit Of Detection (Lod) and Limit Of Quantific...mentioning

confidence: 99%

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DNA extraction procedures and validation parameters of a real-time PCR method to control milk containing only A2 β-casein

et al. 2022

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Section: Dna Isolationmentioning

confidence: 99%

Section: Sensitivity: Limit Of Detection (Lod) and Limit Of Quantific...mentioning

confidence: 99%

DNA extraction procedures and validation parameters of a real-time PCR method to control milk containing only A2 β-casein

et al. 2022

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“…Sequencing is conducted in some cases, but regardless of its efficiency, at this stage, it is not always reasonable for routine large-scale population studies (Dai et al, 2016). The genotyping of cattle by alleles A 1 and A 2 at the beta-casein locus is conducted in different regions of the world which demonstrates the increasing interest in this problem (Zepeda-Batista et al, 2015;Vougiouklaki et al, 2020;Ivankovi´c et al, 2021). In recent years, the interest in genotyping cattle from different breeds has been noted in Ukraine, which is confirmed by a number of publications (Ladyka et al, 2020;Ladyka et al, 2021).…”

Section: Introductionmentioning

confidence: 99%

Comparative analysis of A1 and A2 allele detection efficiency for bovine CSN2 gene by AS-PCR methods

2023

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This study was aimed at conducting a comparative analysis of the efficiency in genotyping cattle by beta-casein locus using the allele-specific PCR methods. The results of the study have demonstrated the necessity to optimize the protocol for the use of AS-PCR to detect alleles A1 and A2. It was found that the use of non-optimized PCR protocols led to the genotyping errors, manifested regarding beta-casein locus (CSN2). The impossibility of using the touchdown PCR as an optimization instrument for AS-PCR was proven. The elaborated typing protocols were used to study the genetic structure of the cattle populations of different breeds, reared in Ukraine – Ukrainian Red-and-White dairy, Ukrainian Black-and-White dairy (two populations), and Charolais. It was found that locus CSN2 was polymorphic in all the cattle populations. The frequencies of allele A2 varied within 0.34–0.91 depending on the population of the animals, which may be conditioned by the specificities in the selection work. No deviation from the Hardy-Weinberg equilibrium was found in any investigated population of cattle.

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Analysis of genotyping features of bovine cattle individuals at the CSN2 locus using ACRS-PCR methods

Kulibaba¹,

Sakhatskyi²,

Liashenko³

2023

Animal Science and Food Technology

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In the context of solving the problem of obtaining high quality dairy products from livestock, the issue of determining the type of beta-casein (A1 and A2) in the protein fraction of milk becomes essential. Purpose – to analyse the use of ACRS-PCR methods for differentiation of A1 and A2 alleles of bovine beta-casein locus. Genotyping features were analysed using the artificially created restriction site polymerase chain reaction method utilising TaqI and DdeI restriction endonucleases. The electrophoretic distribution of DNA fragments in agarose gels of various concentrations was used to analyse restriction patterns. Based on the results of bioinformatic analysis of the nucleotide reference sequences of the experimental fragment of the beta-casein gene, it was found that the primer system for the ACRS-PCR DdeI method is characterised by higher parameters of flanking efficiency of the target DNA site compared to the ACRS-PCR TaqI system due to significantly greater effectiveness of hybridisation of oligonucleotides on the target DNA. Based on the results of laboratory tests of both methods, it is proposed to use an additional procedure for analysing the fluorescence intensity of individual elements of restriction patterns, which allows reducing the number of false genotyping that occurs in both cases (based on the results of using both methods) due to the appearance of non-specific amplification/restriction fragments within the size of target restrictions. The application of the ACRS-PCR DdeI method provides more differentiated patterns of the corresponding genotypes in agarose gel compared to the ACRS-PCR TaqI method, but leads to higher material costs for conducting research. These disadvantages of using primer systems for ACRS-PCR of the beta-casein locus determine the relevance of developing alternative methods for typing A1 and A2 alleles which include allele-specific PCR. The use of results is promising for solving the problems of genotyping cattle individuals of different breeds by A1 and A2 alleles of the beta-casein locus

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Identification of Polymorphisms of Gene CSN2 of B Casein in Greek Cow Breeds (Holstein) by Restriction Fragment Length Polymorphism

Cited by 4 publications

References 21 publications

DNA extraction procedures and validation parameters of a real-time PCR method to control milk containing only A2 β-casein

DNA extraction procedures and validation parameters of a real-time PCR method to control milk containing only A2 β-casein

Comparative analysis of A1 and A2 allele detection efficiency for bovine CSN2 gene by AS-PCR methods

Analysis of genotyping features of bovine cattle individuals at the CSN2 locus using ACRS-PCR methods

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