پژوهشهای علوم دامی ایران (Oct 2022)
Expression of Serine Biosynthesis Pathway Genes in Breast Muscles of Iranian Native Turkeys with Divergent Feed Efficiency
Abstract
Introduction: Feed is the main cost part of poultry production. High feed efficiency poultry produce less feed and less excrement per unit weight gain. Therefore, a comprehensive understanding of the biological mechanisms that control feed efficiency is crucial for the development of optimal breeding and selection strategies. The serine biosynthesis pathway is one of the most important pathways in animals with high feed efficiency. The aim of this study was to investigate the expression of PHGDH, PSAT1 and PSPH genes by real-time PCR in Iranian native turkeys with high and low feed efficiencies.Materials and Methods: Iranian native male turkeys (n=500) were reared up to 20 weeks of age under standard production guidelines. Then 75 turkeys were randomly selected and placed in separate cages with free access to water and feed from 20 to 24 weeks. Turkeys were ranked based on feed conversion ratio (FCR) and three turkeys with the highest and three turkeys with lowest feed efficiency were selected as high feed efficiency (HFE) and low feed efficiency (LFE) birds, respectively. After slaughter of turkeys, RNA was extracted from breast tissue. Quantity and purity of the extracted RNAs were determined using a nanodrop device and its quality was evaluated using 1% agarose gel electrophoresis. Sequences of PSPH, PHGDH, PSAT1 and RSP7 genes were collected from the NCBI database. The primer was designed using Primer Premier version 5 software. All primers were synthesized by Sinaclon (Iran). In this study, RSP7 gene was used as a reference gene. Then, cDNA synthesis was performed. The best amplification temperature for simultaneous amplification of target and reference genes was determined. Samples were amplified for each gene with 3 replications using real-time PCR reaction. Significance level between treatments for each gene was determined separately using t-test in SAS software version 9.2 (P<0.05).Results and Discussion: Results of ultraviolet light absorption measurements at 260 and 280 nm by the nanodrop device showed that the quantity and quality of RNA extracted from the breast muscle samples were of high purity and not contaminated. The range of RNA concentration of the extracted samples was between 480 to 962 ng/μl and the ratio of absorption at 260 and 280 wavelength was about 2.1, which indicates the good quality of the extracted RNAs. The most suitable temperature was selected for specific binding of primers and simultaneous amplification of target genes and temperature control of 58 °C. To investigate and confirm the specificity of replication, melting curves were created to ensure the specificity of the amplified products, the absence of non-specific bands and secondary structures such as hairpin and primer-dimer structures. The results showed that there was only one narrow peak for each gene. The results of studying the expression of serine biosynthesis pathway genes (PSPH, PHGDH and PSAT1) showed that the expression level of these genes in HFE male turkeys was significantly higher than LFE male. Higher expression of PSPH, PHGDH and PSAT1 genes in HFE animals than in LFE animals indicates activation of the serine amino acid biosynthesis pathway, which itself can provide precursors for the Krebs cycle and purine biosynthesis. Glucose is the main source of metabolic energy in the body. When glucose enters the cell, glycolysis begins in the cytoplasm. The pathway of glycolysis and Glutamine catabolism produces an intermediate metabolite called 3-phosphoglycerate, which is gradually catalyzed to serine by PHGDH, PSAT1, and PSPH. Eventually serine is converted to glycine. Activation of this pathway indicates the higher ability of HFE animals to make better use of energy sources such as glucose, which increases protein production in breast muscle tissue and enhances volume and weight of muscle tissue in HFE turkeys.Conclusion: The results of this study showed that the expression of serine biosynthesis pathway genes (PSPH, PHGDH and PSAT1) was significantly higher in high feed efficiency turkeys than in low feed efficiency turkeys. In fact, these results at the level of molecular biology show that turkeys with higher feed efficiency cultivate better use of energy received from feed. Activation of this pathway increases the biosynthesis of various amino acids and thus increases protein and muscle mass in birds. The results of this study can be a promising window to introduce genes that affect feed efficiency in order to further investigate the population and larger flocks of birds.
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