بررسی چندشکلی ژن STAT5B و ارتباط آن با صفات تولیدی در مرغ بومی فارس

Introduction
Genetic resources are one of the most valuable assets of any country. Due to adaptation to specific environmental conditions, Iranian indigenous chickens exhibit remarkable genetic diversity. Identifying and characterizing the genetic makeup of native chicken breeds can facilitate the development of effective conservation and breeding programs. Polygenic traits such as growth and reproduction are economically important in the poultry industry. Single nucleotide polymorphism (SNP) association studies explore the relationship between genetic variants and traits by targeting genes involved in physiological pathways. Signal transducers and activators of transcription (STAT) proteins are a family of cytoplasmic proteins critical in the signal transduction pathways of multiple cytokines and peptide hormones. Several functional polymorphisms in the STAT5B gene have been reported in chickens. Previous studies demonstrated the pivotal roles of STAT5A and STAT5B in physiological processes linked to growth hormone and prolactin in mice. Members of the STAT protein family are involved in growth and reproduction. Identifying genetic markers in candidate genes and their association with economically important traits can optimize breeding programs through marker-assisted selection (MAS). The main goal of this study was to investigate SNP (G4533815A/G-250A) polymorphism in STAT5B gene and its association with production traits in Fars native fowl.

Material and methods
Blood samples were randomly collected from 313 native chickens at the Native Chicken Breeding Farm in Fars Province (Shamsabad station). DNA was extracted using the salting-out protocol and the evaluated through 2% agarose gel electrophoresis and Nanodrop analysis. A 554 bp fragment of the STAT5B gene containing the SNP (G4533815A/G-250A) was amplified using the following primers: F: 5'-CCATCCCTTCCTGGTGCAGT-3' and R: 5'-ACTGCTGCCATTTCCCTTTG-3' with annealing temperature at 60 degrees Celsius. Genotyping was performed using the MspI restriction enzyme at 37 degrees Celsius for 3 hours. Allelic and genotypic frequencies, Hardy-Weinberg equilibrium (HWE), polymorphism information content (PIC) and genetic parameters were calculated using PopGene3.2 software. The distribution of phenotypic data was not normal; therefore, nonparametric statistics were used to analyze the data. The association between the STAT5B gene polymorphism and production traits (egg number [EN], age at first egg [AFE], egg weight at 84 weeks [EW84], and body weight at 12 weeks [BW12]) was analyzed using the Mann-Whitney test in Minitab (version 18). Gene networks associated with the STAT5B gene in Gallus gallus domesticus were visualized using the STRING web tool (version 12).



Results and discussion
SNP genotyping identified three genotypes: AA (128 individuals, uncut fragment - no mutation), AG (174 individuals, heterozygote, 554/477 and 77 base pairs fragments - one mutant allele), and GG (11 individuals, 477 and 77 base pairs fragments - two mutant alleles). The frequencies of the A and G alleles were 0.69 and 0.31, respectively. The chi-square test (ꭓ² = 31.47) indicated that the population deviated significantly from HWE (P < 0.01). This deviation can be attributed to selection intensity and mating systems. The Mann-Whitney test revealed a significant difference in AFE between genotypes AA/GG (W = 191.50) and GG/GA (W = 354.50). The GG genotype had a lower median AFE (142 days) compared to AA (150 days) and AG (147 days). Previous studies have demonstrated that hormones, cytokines, and their receptors regulate growth, reproduction, immunity, and metabolism in animals. The chicken STAT5B gene has been implicated in sexual maturation via ovary development regulation. Protein interaction network analysis also revealed that STAT5B interacts with proteins related to growth, immunity, and reproduction, reinforcing its importance in controlling production and economic traits.
Conclusions
The results of the present study showed that there is a significant association between the SNP (G4533815A/G-250A) polymorphism of STAT5B gene and age at first egg, but no significant association was reported with other traits such as egg weight, egg number, and body weight. The main reason for the difference between the provided results and other investigations can be attributed to the statistical method used and the different genetic potential of the populations. Identification of beneficial mutations and genetic pathways affecting reproductive traits such as age at first gg of chickens can be used to select the desired allele in the population in order to achieve appropriate performance. Identification of candidate genes affecting economic traits can be used in marker-assisted selection breeding programs. The findings provide applied strategy to improve Fars native chicken breeding programs.