| چکیده انگلیسی مقاله |
Extended abstract
Introduction and Objective: FEZL is one of the candidate genes related to performance, quality traits of milk and resistance to mastitis disease in dairy cows. Mastitis is one of the common diseases in the dairy cattle population, which imposes a lot of economic losses on the farmer. In the studies that were conducted in order to investigate the genetic variation in the FEZL gene locus and their relationship with the milk somatic cell count (SCC), it was shown that there is a significant relationship between the genetic variants in the mentioned locus and the milk somatic cell count, which is indicative of mastitis. Also, conflicting results of the effects of the mentioned gene variants on the quantitative and qualitative traits of milk have been reported. Therefore, the purpose of this research is to investigate the amount of different deletion, addition and mutation polymorphisms in the FEZL locus, to identify important variants related to milk composition, and to investigate the difference between pure breeds and their crossbreds of Sistani and Holstein cows.
Materials and Methods: In this research, 20 pure breeds of Sistani and Holstein cows (10 cows from each group) and their crossbreds of Sistani and Holstein (7 heads) from the Sistani cattle research center and native herds of Sistan region. Blood was randomly selected and taken from the jugular vein. Quantitative and qualitative traits of milk were measured in a period of three months from morning and evening milking. Extraction of total DNA from blood samples was done by salt-optimized method. Then, the extracted DNA was electrophoresed on a 0.5% agarose gel and the samples that were of good quality were used for the PCR amplification reaction. To amplify the 229 bp fragment from the FEZL gene locus, a specific primer pair was used with temperature cycling in a thermocycler. Sequencing was done with Chromas software in FASTA format and low-quality sequences were removed. The next step is to remove the production areas of a single thread from the back-and-forth sequence, canting production was done using CAP3 software. Alignment of 27 samples was done with MEGA software in pairs and multiples. 4 samples were removed due to unfavorable alignment and 23 samples were saved for further analysis. The analysis of polymorphic regions caused by mutations, deletions and additions and haplotyping was done with MEGA6 and DNASP5 software, and finally, the effect of each marker with milk quality traits was estimated using the single marker model and GLM procedure of SAS9 software. To reduce the type 1 error in multiple tests, the Benferroni test was used at the 5% probability level. Finally, comparing the mean of the groups with the Tukey-Kramer method was used at the probability level of 5%.
Results: A total of 86 polymorphic regions were identified in exon 1 of FEZL gene among pure and crossbred populations. In all samples, the percentage of transitional substitution was 32.56% and transversional substitution was 67.44%. The percentage of transitional and transversional substitutions were estimated as 50.6 and 49.4% in Holstein breed, 28.08 and 71.92% in Sistani breed and 33.55 and 66.45% in crossbreds, respectively. By comparison, the percentage of transitional substitution was the lowest in Sistani breed and the highest in Holstein, but the percentage of transversional substitution was the lowest in Holstein and the highest in Sistani. The coefficient of transition substitution ratio to transversional, which is an indicator of evolution during the past periods, was the highest in Holstein (0.75) and the lowest in Sistani (0.30). In all populations, the frequency of bases in the nucleotide sequence of the exon1 region of FEZL gene, Thymine was the lowest and Cytosine was the highest frequency. The highest diversity within breeds was related to Holstein and Sistani crossbreds (0.15) and the lowest diversity was for Sistani breed (0.044). From the degree of divergence among different breeds, the highest degree of divergence was between crossbreds with Holstein (0.126) and the lowest between Sistani and crossbreds (0.012). Of the 77 polymorphic loci (SNP), 33 cases were two-variant, 9 cases were three-variant, and two were four-variant. In the milk composition, the percentage of fat, protein, lactose and fat-free dry matter was the highest in the Sistani breed and the lowest in the Holstein breed, and it was in the middle in the crossbreds. The effect of breed on all milk traits except protein percentage was significant (P<0.01). Among the identified markers, the marker at the position of 97 bp of the exon1 of FEZL gene had the highest effect on the traits of fat, lactose, protein and milk dry matter percentage (P<0.01).
Conclusion: despite the existence of different polymorphisms in the exon1 of a FEZL gene and the amount of genetic diversity that exists between and within the Holstein and Sistani breeds, it can be considered as a useful marker in the breeding of native livestock and from the strength The combinability of variants in crossbreds was used to improve the economic characteristics of milk. Also, polymorphism in the mentioned area was still used as an indicator to improve somatic cell counts in milk as an indicator of mastitis control in dairy cows, which was not possible to measure in this research and is recommended in future studies.
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