Signatures of selection in the genome of Tsarskoye Selo chicken population emerging as an individual breed

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Introduction

Runs of homozygosity (ROH) are a crucial tool for detecting signatures of selection, studying genetic diversity, tracing breed formation history, and understanding mechanisms of selection and inbreeding. These regions represent clusters of DNA where individuals carry identical homozygous genotypes, indicating the influence of factors such as selective breeding or inbreeding. Numerous chicken breeds have been analyzed through ROH, revealing selection signatures linked to genes associated with traits targeted by breeding programs [1]. For newly developed breeds, such as Tsarskoye Selo, ROH analysis enables the identification of selection footprints within gene loci subjected to selective pressure, as well as assessment of inbreeding levels and breed formation dynamics.

The creation concept of Tsarskoye Selo chicken breed was based on breeding a large meat-egg autosexing breed characterised by accelerated growth rates in juvenile birds, substantial egg mass and notable adaptability to diverse housing environments. Poultry breeders began work on breeding in 1991 at the Russian Research Institute of Farm Animal Genetics and Breeding. The birds were named ‘Tsarskoye Selo’ in honour of the imperial residence Tsarskoye Selo, referred to today as Pushkin, where the work on breeding these chickens took place. Among scientific studies, derivative forms of the name ‘Tsarskoye Selo’ may also be found: Tsarskoselskaya, Tsarskoselsky and similar.

Selection of Tsarskoye Selo chickens were conducted by meat-purpose breeds such as Cornish and White Plymouth Rock to produce four-line rooster hybrids that then were mated with dual-purpose breeds New Hampshire and Poltava Clay [2]. Cornish and White Plymouth Rock set the meat direction for Tsarskoye Selo. Cornish, known as Indian Game or Cornwall, are valued for their large amount of breast meat, in which they are unrivalled among many other breeds. Traditionally, Indian Game males were selected as partners for females of other meat breeds to produce large meat birds and broilers. Thus, a creation of meat hybrid, which is a progenitor of the initial Tsarskoye Selo population, was provided by mating Cornish roosters with White Plymouth Rock hens, which are also esteemed for their meat qualities. Moreover, Plymouth Rock and Cornish have retained the egg-brooding instinct characteristic of heavy breeds, which Tsarskoye Selo inherited from them while maintaining an excellent egg-laying quality. [3], [4]. The females New Hampshire and Poltava Clay set the meat and egg direction, and endowed Tsarskoye Selo with relative early maturity and adaptive properties, given that New Hampshire and Poltava Clay reach sexual maturity early, are hardy, and easily acclimatize to adverse conditions. [3], [5]. Poultry farmers note that Tsarskoye Selo chickens are stress-resistant, they well-tolerate to withstand winter conditions in domestic poultry houses, and compose demeanour in flocks in comparison with other breeds as Leghorn, Italian, and Czech. Tsarskoye Selo are characterised by high levels of mobility, driven by an aptitude to spacious placement; however, they are also content with compact space. They possess a calm temperament, which is indicative of strong animals; besides, they get along well with humans, and are amenable to taming.

A significant task for breeders was to consolidate morphological traits associated with plumage colouration. A new breed must possess specific genetic markers enable the differentiation between cockerels and pullets at the day-old chicks, along with distinctive color patterns attributable to ancestral hereditary complex. For this purpose, the solution was the development of a genetic system that includes targeted alleles for chick sexing. The paternal lines of the four-line hybrids carried the target alleles dominant wheaten (e^Wh) and brown (e^b) that are likewise the non-target alleles extended black (E), recessive wheaten (e^y), birchen (E^R) and wild type (e⁺) have different phenotypic effects on black color manifestation and belong to the MC1R Melanocortin 1 Receptor gene, localized on chromosome 11 [6], [7]. Other target alleles for sexing chickens that were present in the meat hybrid parents affect the expression of gold (s) or silver (S) colors and are associated with the Z chromosome in birds. Dominant homozygous alleles B/B (barring) in meat hybrids, which have an important selective purpose, cause a barred feather pattern and are associated with two non-coding and two coding mutations in the locus of the CDKN2A - cyclin-dependent kinase inhibitor 2A. CDKN2A is localized on the Z chromosome and has a greater phenotypic effect in roosters due to male homozygosity for the Z chromosome and the resulting codominance. [8], [9], [10], [11]. The New Hampshire and Poltava Clay maternal lines had one recessive barring allele, a recessive golden (s) allele, and two wheaten alleles (e^Wh). When the first generation was obtained, they differ in genotypes and phenotypes; hence, futher selection was carried out according to the principle of selecting buff barred individuals. As the result of staged hybrid selection and back-crossing with New Hampshire breed, the current Tsarskoye Selo chicken population is consolidated with and distinctive buff barred plumage (figure 1) and  autosexing genetic markers (table 1). More detailed information about the stages of selection work and the algorithm for transferring target alleles of feather coloring genes can be found in Makarova AV dissertation^[1] and researches with co-authors Jurchenko OP, Vakhrameev AB [13], [12], [14].

Figure 1. External appearance of Tsarskoye Selo chickens. On the left side – male, right side - female. A buff-barred large fowl with a predominance of yellow shades in plumage.

More than 30 years have passed since the origin of Tsarskoye Selo. At present, their external characteristics correspond to those of heavy birds with superior meat quality; additionally, their latitudinal body proportionality index supports their classification as a dual-purpose breed [15].  Evaluations of both meat and egg quality substantiate the dual-purpose potential of the Tsarskoye Selo chickens within the poultry industry. Herewith, selective breeding aimed at improving meat quality continues, with observable progress in increasing body weight across generations in both roosters and hens. [16].

Table 1. Adapted composite crossbreeding scheme comes from studies [12], [14].

Z и W - sex chromosomes in chicken. When ZW are not specified in genotype entry, alleles are assumed autosomal. B – barring, sex-linked dominant allele; b – barring , sex-linked recessive allele; S – silver feather colour, sex-linked dominant allele; s - gold feather colour, sex-linked recessive allele. E – black plumage, e^Wh  - dominant wheaten, e^b – brown. Co/co - Columbian coloration limits the distribution of black coloration to the flight feathers, mane feathers and tail feathers.

Mating №	Stages	PARENTS		PROGENY
		♀	♂
1	Initial crossbreeding	New Hampshire, Poltava Clay	Cornish and White Plymouth Rock meat hybrid	1st generation hybrids
		ZbW ZsW  eWh/eWh Co/Co	(ZBZb + ZBZB) ZSZs EWh/eb co/co	(ZBZb + ZBW) (ZSZs + ZsW) (eWheWh + E eWh) Co/co
2-5	Mating hybrids with buff barred plumage	1st generation hybrids, then following generations	1st generation hybrids, then following generations	2nd to 5th generations
		ZBW ZsW eWh/eWh Co/co	(ZBZB + ZBZb) ZsZs eWh/eWh Co/co	(ZBZB + ZBW) (ZsZs + ZsW) eWh/eWh (Co/Co + Co/co + co/co)
6	Mating homozygous to secure traits of interest in next generation	5th generation	5th generation	6th generation
		ZBW ZsW  eWh/eWh Co/Co	ZBZB ZsZs eWh/eWh Co/Co	(ZBZB + ZBW) (ZsZs + ZsW) eWh/eWh Co/Co
7	Backcross with New Hampshire	New Hampshire	6th generation	7th generation
		ZbW ZsW eWh/eWh Co/Co	ZBZB ZsZs eWh/eWh Co/Co	(ZBZb + ZBW)  (ZsZs + ZsW) eWh/eWh Co/Co
8	Mating obtained females with males of previous generation	7th generation	6th generation	Next generation
		ZBW ZsW eWh/eWh Co/Co	ZBZB ZsZs eWh/eWh Co/Co	(ZBZB + ZBW) (ZsZs + ZsW) eWh/eWh Co/Co

The productivity of the Tsarskoye Selo population may have been impacted by genetic characteristics that became established during the selection process. In essence, target features are predominantly located within regions of homozygosity. They facilitate the identification of associations with phenotype and the estimation of inbreeding extent. A recent study of the genetic divergence among various chicken breeds [17] revealed that Tsarskoye Selo stood out as a separate cluster, but retained a close genetic distance to Poltava Clay and New Hampshire, which is consistent with the data of phylogenetic analysis using Neighbor-Net, showing how the Tsarskoye Selo population branches off from the maternal population of the Poltava Clay breed [18]. At the same time, PC analysis demonstrates a strong divergence of Tsarskoye Selo from the Cornish and White Plymouth Rock breeds [17].

The objective of this study is to identify homozygous regions in the Tsarskoye Selo chicken population and to analyse the linked group of genes concentrated in the ROHs found. The results of the study will provide valuable insights into the role of homozygous regions in enhancing the productive qualities, external characteristics, and adaptive properties of the Tsarskoye Selo population.

Materials and methods

A total of 12 chickens of Tsarskoye Selo population were provided by Department of bioresource collection of Russian Research Institute of Farm Animal Genetics and Breeding (Puskin, Russia). A total of 1 ml blood from the wing vein of each individual was collected in EDTA tubes and stored at -20°C for subsequent DNA extraction. Genomic DNA was extracted using the phenol method. The concentration and quality of the samples with extracted DNA solution was determined on a NanoDrop2000c spectrophotometer (ThermoFisher Scientific, USA). The concentration of each sample averaged 400 ng/μl, and the purity of the samples at an optical density ratio of A260/280 was greater than 1.9. All samples underwent quality control and were sequenced with 30x coverage. Libraries were prepared using the TruSeq DNA Nano Kit (Illumina Inc., San Diego, California, USA). Sequencing was performed using NovaSeq 6000 (Illumina Inc., San Diego, California, USA) with a read length of 2 × 151 bp. Quality control showed that 91.46% of reads met the Q30 threshold. Read quality assessment and filtering were performed using FastQ v 0.12.0. The obtained reads were aligned to the reference genome of the red jungle fowl Gallus_gallus_gca000002315v5.GRCg6a from the international ENSEMBL database using the bwa-mem2 software package for mapping short reads to large reference genomes. As a result, a total of 20,077,197 SNPs were detected. SNP filtering was performed using PLINK 1.9 with the following parameters: --maf 0.05, --geno 0.02, and --hwa 0.0001, resulting in 11,788,716 SNPs remaining in the analysis. SNPs on sex chromosomes were excluded from the analysis to eliminate the influence of sex on the assessment.  ROH regions were analysed using the detectRuns package in the RStudio v 2023.12.1+402 software environment, with the following parameters applied: a window size of 150 SNPs; a window overlap threshold of 0.1; a minimum number of SNPs in a region of 200; and a maximum number of heterozygous SNPs in the window of 1. Then, for the detected ROH regions, boundaries corresponding to the GRCg6a reference genome assembly were determined. To identify promising candidate genes, the annotation of genes located in these regions was performed in the Ensembl genome browser.

Results

Identification of ROH Islands and Gene Annotation

The sample we studied showed variability in the number of homozygous regions between individuals, indicating genetic diversity and differences in genetic background; nevertheless, ROH occurred in more than 75% of individuals on chromosomes 5 and 11 (Figures 2 and 3).

Figure 2. Distribution of homozygous regions (ROH) on GGA5 for each individual.

 

Figure 3. Distribution of homozygous regions (ROH) on GGA11 for each individual.

At the same time, we discovered a completely conservative and homogeneous homozygous region on chromosomes 5 and 11 in all studied individuals, indicating the potential functional significance or evolutionary conservatism of this genome fragment (Figures 4 and 5).

Figure 4. Distribution of homozygous regions (ROH) on GGA5. The X-axis represents the chromosome up to 60 Mbps, the Y-axis represents the percentage of homozygous regions, and Ts represents the Tsarskoye Selo chicken population.

Figure 5. Distribution of homozygous regions (ROH) on GGA11. The X-axis represents the chromosome up to 20 Mbps, the Y-axis represents the percentage of homozygous regions, and Ts represents the Tsarskoye Selo chicken population.

During the analysis of ROH regions, the target genes were annotated (table 2). The annotation of target genes provided the functionality and biological significance of these regions, as well as the identification of genes associated with the phenotypes or adaptive traits of interest.

Table 2. Tsarskoye Selo ROH regions; gene locations according to the Chicken genome assembly - (Red Jungle fowl), GRCg6a, INSDC Assembly GCA_000002315.5, (March 2018); and biological functions of noted genes.

Chr	Length of ROH	Start, bp	End,bp	Annotated genes	Biological process
GGA5	40,718,607 - 41,100,083	40,756,879	40,852,483	DIO2	Thyroid hormones biosynthesis
		40,872,467	40,967,218	CEP128	Biogenesis of structures associated with cell division
		40,973,189	41,021,853	TSHR	Thyroid cell metabolism control
		41,030,173	41,055,257	GTF2A1L	RNA polymerase II pre-initiation complex assembly
		41,074,005	41,141,503	STON2	Endocytotic complexes regulation
GGA11	17,765,088 - 17,960,178	17,767,709	17,789,534	FBXO31	Participates in proteolysis pathways
		17,796,398	17,805,107	MAP1L1C3B	Regulating autophagy of intermediate filaments in cells
		17,807,235	17,852,384	ZCCHC14	Peroxisome regulation
		17,866,029	17,902,011	JPH3	Endoplasmic reticulum components regulation
		17,911,374	17,940,151	KLHDC4	Ubiquitination substrates control
		17,955,150	17,987,235	SLC7A5	Cationic transport of amino acids associated with glycoprotein

In the first region, we selected candidate genes from those annotated in the Ensembl database that are associated with chicken productivity: DIO2 (deiodinase, iodothyronine type II), CEP128 (centrosomal protein 128), TSHR (thyroid stimulating hormone receptor), GTF2A1L (general transcription factor IIA subunit 1), STON2 (stonin 2).

In the second homozygous region, we selected target genes associated with adaptive traits in chickens: FBXO31 (F-box protein 31), MAP1L1C3B (microtubule associated protein 1 light, ZCCHC14 (zinc finger CCHC-type containing 14), JPH3 (junctophilin 3), KLHDC4 (kelch domain containing 4), SLC7A5 (solute carrier family 7 member 5).

Conclusion

GGA5 ROH Gene Functions

When analysing the homozygous region on GGA5, a section with linked genes was discovered: DIO2, CEP128, TSHR, GTF2A1L, STON2. A recent study by Basheer, A., Haley, C.S., 2015 notes strong selective pressure on this locus and emphasises its regulatory effect on the thyroid-pituitary axis. [19]. Hence, this linked group of genes is of great practical interest, as they act on the hormones of the thyrotropic axis, which in turn are involved in many processes throughout the life of chickens. During embryogenesis, thyroid hormones transferred from the mother's body to the embryo coordinate gene expression, thereby ensuring the correct sequence of morphogenesis of tissue and organ cells. [20]. In the context of ontogenesis, thyroid hormones fulfill a role in metabolic processes and the formation of productive and reproductive qualities. They ensure normal energy metabolism by increasing the number of mitochondria, energy and heat production, and tissue oxygen demand. In coordination with metabolic processes, the thyroid profile stimulates folliculogenesis, maintains egg production in hens, and determines 'brooding' qualities. [21], [22], [23], [24], [25].

The thyroid stimulating hormone receptor has several protein products as a result of alternative splicing, which are expressed depending on the alternatively spliced variants of TSHRa-f either in the thyroid gland or in the chicken pituitary gland. [26], [27]. A series of studies devoted to the history of chicken breeding and the study of the genotype-phenotype relationship with regard to genes under selection pressure has shown the key role of TSHR in the domestication of chickens. [25], [28], [29]. It is noteworthy that researchers at the L.K. Ernst Institute of Animal Husbandry discovered the same ROH on GGA5 in Cornish as we found in Tsarskoye Selo in our study. [30]. The transmission of this short homozygous region from Cornish suggests that there has been long-standing inbreeding, and identifies genes that have undergone natural or artificial selection in past adaptation and breeding processes.

In birds, photoreceptors located deep within the brain mediate the photoperiodic regulation of gonadal responses. This regulatory mechanism involves the transformation of TSH receptor proteins facilitated by the activity of the DIO-2 gene [31]. DIO2 exists in birds alongside other deiodinases, DIO1 and DIO3, which are homologous to their human counterparts. The primary role of deiodinases is to catalyze the conversion of thyroxine (T4), which circulates in the bloodstream in an inactive form, into the active hormone triiodothyronine (T3) within tissue and organ cells. DIO1, DIO2, and DIO3 differ in their tissue-specific expression and their distinct roles in the removal of iodine atoms either from the phenolic outer ring (5′ deiodination or ORD) or the tyrosyl inner ring (5 deiodination or IRD). The non-selective DIO1 catalyzes both ORD and IRD, DIO3 exclusively catalyzes IRD, while DIO2 specifically catalyzes ORD [23]. DIO2 is predominantly expressed in the brain, where it plays a crucial role in the local conversion of thyroxine (T4) to triiodothyronine (T3). In precocial birds - those that hatch fully mobile - there is an increased concentration of T3 in the brain at hatching due to enhanced local conversion of T4 to T3 mediated by DIO2 [32].

A study by Deng Y, et al., 2021 [33] revealed the key role of the DIO2 gene in the formation of bony structure, known as protuberant knob, observed in certain domestic goose breeds. Histomorphological analysis showed that this knob consists of a pneumatized bone structure. In geese, the protuberant knob serves as an indicator of sexual maturity [34].

The CEP128 gene encodes a protein belonging to the family of centrosomal proteins. These proteins are active components in the biogenesis of centrioles and the centrosome, playing crucial roles in cell division and cell cycle regulation. They are involved in spindle-kinetochore assembly, centriole duplication, cell polarity, and the signaling mechanisms of cell cycle checkpoints [35]. Notably, CEP128 appears to have no obvious linkage disequilibrium with TSHR. However, SNPs within CEP128 are associated with a genetic predisposition to thyroid diseases. Furthermore, gene set enrichment analysis (GSEA) has linked CEP128 to several common immune pathways, including the interferon-γ signaling pathway and the toll-like receptor signaling pathway, both of which play roles in the pathogenesis of Graves’ disease designated as autoimmune thyroid disorder [36].

The genes GTF2A1L and STON2, identified in regions of homozygosity (ROH), are potentially linked to egg production. GTF2A1L encodes the TFIIA alpha and beta-like factor, which is involved in the assembly and stability of the RNA polymerase II pre-initiation transcription complex. This gene is specific to germ cells and may play a critical role in testicular biology and spermatogenesis. Certain polymorphisms and mutations in GTF2A1L have been associated with human disorders such as Leydig cell hypoplasia type 1, peripheral precocious puberty, and infertility [37], [38]. Meanwhile, STON2 has been identified as a candidate gene influencing egg-laying performance in chickens [39]. Through neural networks, STON2's function is linked to folliculogenesis and yolk weight regulation [40]. 

GGA11 ROH Gene Functions

In study Sun Y. et al., 2023 was identified a genomic region linked to feather traits in ducks that includes the same genes we found in a conserved ROH region on GGA11. Both significant and suggestive SNPs were detected in the genes ZCCHC14, KLHDC4, and SLC7A5, which showed associations with distinct feather pigmentation patterns [41].

The identified gene FBXO31 encodes a protein that is one of the four subunits of the SCF (SKP1-Cullin-F-box) ubiquitin-protein ligase complex, which is involved in proteolysis pathways. Studies in chickens indicate that FBXO31 functions to halt the cell cycle and respond to DNA damage, thereby reducing the risk of cancer development. Molecular evolutionary analysis of the GOLPH3 gene repertoire has revealed a mechanism involving FBXO31 that contributes to birds' lower susceptibility to cancer compared to mammals [42], [43].  

The gene KLHDC4, which belongs to the conserved Kelch superfamily and the KLHDC subfamily, performs an ubiquitination function. Kelch domains form scaffold structures that facilitate protein interactions by binding substrates for ubiquitination and subsequent degradation. Kelch proteins localize in the cell in ways that suggest roles in DNA and RNA biological processes. KLHDC4 specifically localizes to nucleoli and kinetochores and shows high expression levels in the endometrium and mammary gland, highlighting its potential involvement in cell division and meiotic recombination [44], [45]. Ubiquitin tagging of proteins activates the proteasome system, which connects to autophagy processes through shared regulators or substrates. The gene MAP1LC3B serves as a key marker for autophagy and allows tracking of this process in cells. It shows high expression in primordial germ cells (PGC). The MAP1LC3B protein regulates autophagy during cell differentiation and promotes the accumulation of autolysosomes, which break down intermediate components during the formation of primary protective cells [46], [47]. Oxidative stress triggers increased glucocorticoid production, which places higher demands on the hypothalamic-pituitary-adrenal axis. In response, cells upregulate MAP1LC3B expression, directing it toward lipid metabolism and autophagy [48].

The function of the ZCCHC14 gene was determined in a study of antiviral responses to Sendai virus using an optical pool-screening method combined with a genome-wide analysis of hidden morphological traits. In mammals, ZCCHC14 acts as a regulator of peroxisomes, reducing peroxisomal activity and participating in cellular antiviral defenses [49]. At the protein level, ZCCHC14 functions as an adaptor for terminal nucleotidyltransferase 4A (TENT4A or PAPD7), enhancing cellular mRNA stability through non-canonical polyadenylation [50].

We discovered two genes, JPH3 and SLC7A5, in the described ROH region, both linked to membrane transport of specific molecules and compounds. JPH3 belongs to the junctophilin family, which helps establish, maintain, and modulate the structure and function of calcium microdomains - specialized junctions between the plasma membrane and the endoplasmic reticulum. Junctophilins connect the cell surface with intracellular ion channels to trigger and activate calcium-selective channels in the plasma membrane. This mechanism is essential for calcium signaling and initiating muscle fiber contraction and neuron activation [51]. In mammals, JPH3 expresses mainly in the brain. It regulates subsurface cisternae of the endoplasmic reticulum near the plasma membrane. Knockout of JPH3 in mice leads to severe neuromuscular defects, growth retardation, deficits in short- and long-term memory, and lethality by one month of age [51].

SLC7A5 belongs to the SLC (Solute Carrier) superfamily of membrane transport proteins. These proteins transport dissolved substances across cell membranes, regulate acid-base balance, mediate neurotransmission, and support energy metabolism in both anabolic and catabolic processes. The SLC7A5 gene is part of the SLC7 family, which includes cationic amino acid transporters and glycoprotein-associated amino acid transporters [52]. SLC7A5 forms heterodimers with SLC3A2 to activate the mTORC1 signaling pathway and enable type 1 amino acid transport, contributing to cellular resistance against ferroptosis induced by oxidative stress. [53], [54]. In study Khwatenge CN, et al., 2020 were identified several candidate genes from the SLC7 family that influence body weight gain in broiler chickens. Their transcriptomic analysis revealed a strong correlation between lysine intake and body weight gain. Increasing dietary lysine levels led to elevated expression of SLC7 family genes, which in turn stimulated protein synthesis through activation of the mTORC1 pathway [55].

Our study of the annotated genes in this homozygous region links their functions to cellular metabolism, substance transport, and immune response regulation. The formation of this homozygous region likely reflects selective pressure driving genetic predisposition toward enhanced adaptive traits. These adaptations operate through mechanisms involving DNA repair, catabolism of intermediate compounds, and homeostasis maintenance. Together, these processes contribute to stress resilience and a robust immune response.

Regions of homozygosity summary

Our study identified homozygous regions, including two on chromosomes 5 and 11 that are inherited by 100% of the Tsarskoye Selo chickens in the population we examined. The genes located in these homozygous regions contribute to phenotypic traits that distinguish this population as a distinct breed. Analysis of the homozygous region on chromosome GGA5 revealed a significant QTL containing a cluster of linked genes—DIO2, CEP128, TSHR, GTF2A1L, and STON2. These genes play key roles in embryonic development, metabolism, productivity, and reproductive traits, notably through the regulation of thyroid hormones, which coordinate morphogenesis and energy metabolism. The TSHR gene, in particular, has undergone selective pressure during chicken domestication. The presence of the same homozygous region in the Cornish breed confirms a history of inbreeding and highlights genes that have been targets of natural or artificial selection. This linked gene cluster within the homozygous region on GGA5 represents a valuable target for future research and selective breeding, given its impact on thyroid hormone profiles, development, productivity, and adaptation in chickens. This ROH region corresponds with egg production and productivity, which underpinned the creation of the Tsarskoye Selo breed.

The ROH region identified on chromosome 11 in Tsarskoye Selo overlaps with previously described genomic areas in ducks associated with feather pigmentation. Genes in this homozygous segment participate in critical cellular processes, including metabolism, immune response, DNA repair, and homeostasis maintenance, underscoring the region’s importance in adaptation and resilience to stress. Consequently, the linked gene cluster in this ROH region reflects selective pressures aimed at enhancing adaptive traits and immune response through complex molecular mechanisms, a desired outcome in breed development.

ROH regions emerge in populations through intentional inbreeding as well as through crossbreeding individuals from different breeds sharing identical homozygous segments. In the Tsarskoye Selo population, short ROH segments suggest historical inbreeding or genetic drift, whereas the absence of long ROH regions indicates no recent inbreeding.

Discussion

Analysis of the identified runs of homozygosity regions on chromosomes 5 and 11 revealed a concentration of key genes influencing developmental processes, productivity, reproductive traits, and adaptability in poultry. Notably, genes associated with the thyrotropic axis, including TSHR and DIO2 among others, play critical roles in coordinating metabolic regulation, performance, and resilience. This is further supported by the presence of a homologous homozygous region in the Cornish breed, from which the Tsarskoye Selo population inherited part of its genome. The homozygous region on chromosome 11 is linked to stress resistance pathways, DNA repair mechanisms, immune response and feather pigmentation.

These findings collectively demonstrate that the homozygosity pattern observed in the Tsarskoye Selo genome reflects intentional artificial selection aimed at fixing desired productive, adaptive, and morphological traits. The presence of distinct ROH segments underscores the successful formation of these birds, which has inherited essential characteristics from foundational breeds and validates its status as an independent autosexing meat-egg breed. This genomic insight lays a robust foundation for future molecular genetic studies and breeding programs targeted at enhancing productivity and resilience within the breed population.

 

¹ Makarova AV. Gene fund chicken breeds serve as the material for auto-sex populations and productive hybrids. [dissertation].  06.02.07., 21.01.2020., Moscow. EDN: CAPUMH

About the authors

Yuri Sergeevich Shcherbakov

Russian Research Institute of Farm Animal Genetics and Breeding — Branch of the L.K. Ernst Federal Research Center for Animal Husbandry

Email: yura.10.08.94.94@mail.ru
ORCID iD: 0000-0001-6434-6287
SPIN-code: 3547-1009
Scopus Author ID: 57221619264
ResearcherId: AAR-5595-2020

PhD in Biology

Researcher 

Laboratory of molecular genetics

Russian Federation, 196601, St. Petersburg, Pushkin, Moskovskoe shosse, 55a

Olga Anatolevna Nikolaeva

Russian Research Institute of Farm Animal Genetics and Breeding — Branch of the L.K. Ernst Federal Research Center for Animal Husbandry

Author for correspondence.
Email: helgaa.nikolaeva@gmail.com
ORCID iD: 0000-0003-3828-1111
SPIN-code: 7193-6004
Scopus Author ID: 57941963300
ResearcherId: GPK-6027-2022

Junior Researcher 

Laboratory of molecular genetics

Russian Federation, 196601, St. Petersburg, Pushkin, Moskovskoe shosse, 55a

Anatoly Borisovich Vakhrameev

Russian Research Institute of Farm Animal Genetics and Breeding — Branch of the L.K. Ernst Federal Research Center for Animal Husbandry

Email: ab_poultry@mail.ru
ORCID iD: 0000-0001-5166-979X
SPIN-code: 6810-7339
Scopus Author ID: 56862214400
ResearcherId: AAD-1068-2022

Senior Researcher

laboratory for scientific support of the conservation of poultry genetic resources

 

Russian Federation, 196601, St. Petersburg, Pushkin, Moskovskoe shosse, 55

Anastasia Ivanovna Azovtseva

Russian Research Institute of Farm Animal Genetics and Breeding — Branch of the L.K. Ernst Federal Research Center for Animal Husbandry

Email: ase4ica15@mail.ru
ORCID iD: 0000-0002-2963-378X
SPIN-code: 5784-2786
Scopus Author ID: 57942391700
ResearcherId: MSZ-1418-2025

Junior Researcher 

Laboratory of molecular genetics

Russian Federation, 196601, St. Petersburg, Pushkin, Moskovskoe shosse, 55a

Natalia Victorovna Dementieva

Russian Research Institute of Farm Animal Genetics and Breeding — Branch of the L.K. Ernst Federal Research Center for Animal Husbandry

Email: dementevan@mail.ru
ORCID iD: 0000-0003-0210-9344
SPIN-code: 8768-8906
Scopus Author ID: 57189759592
ResearcherId: T-4551-2018

PhD in Biology

Chief Researcher 

Laboratory of Molecular Genetics

196601, St. Petersburg, Pushkin, Moskovskoe shosse, 55a

References

Supplementary files