Association between cholecystokinin type A receptor haplotypes and growth traits in Japanese Hinai-dori crossbred chickens

We previously identified quantitative trait loci for body weight and average daily gain in a common region between MCW0240 (chr 4: 69.9 Mb) and ABR0622 (chr 4: 86.3 Mb) on chicken chromosome 4 in an F₂ resource population produced by crossing low- and high-growth lines of the Hinai-dori breed. Cholecystokinin type A receptor (CCKAR) is a candidate gene affecting growth traits in the region. In this study, we genotyped polymorphisms of the CCKAR gene and investigated its association with growth traits in a Hinai-dori F₂ intercross population. All the exons of the CCKAR gene in the parental population were subjected to PCR amplification, nucleotide sequenced and haplotypes identified. To distinguish resultant diplotype individuals in the F₂ population, a mismatch amplification mutation assay was performed. Five haplotypes (Haplotypes 1–5) were accordingly identified. Six genotypes produced by the combination of three haplotypes (Haplotype 1, 3, and 4) were examined in order to identify associations between CCKAR haplotypes and growth traits. The data indicate that Haplotype 1 was superior to Haplotype 3 and 4 in body weight at 10 and 14 weeks of age, average daily gain between 4 and 10 weeks, 10 and 14 weeks, and 0 and 14 weeks of age. It was concluded that CCKAR is a useful marker of growth traits and could be used to develop strategies for improving growth traits in the Hinai-dori breed.     

Polymorphism of Heart Fatty Acid-Binding Protein Gene Associatied with Fatness Traits in the Chicken

Fatty acid-binding proteins (FABP) belong to a superfamily of lipid binding proteins that exhibit a high affinity for long chain fatty acids and appear to function in metabolism and intracellular transportation of lipids. The current study was designed to investigate the effects of heart (H)-FABP gene on chicken growth and body composition traits. The Northeast Agricultural University divergent broiler lines for abdominal fat and a broiler X silkie F₂ population were used in this study. Body weight and body composition traits were measured in the populations. Primers were designed according to the chicken H-FABP gene sequence. Polymorphisms between parental lines were detected by DNA sequencing. PCR-RFLP and PCR-fragment length polymorphism methods were developed to genotype the populations. The results showed that the H-FABP gene polymorphisms in the two populations were associated with abdominal fat percentage. It implied that H-FABP gene can be a candidate locus or linked to a major gene(s) that affects abdominal fat content in the chicken.     

Two insertion/deletion variants in the promoter region of the QPCTL gene are significantly associated with body weight and carcass traits in chickens

Glutaminyl‐peptide cyclotransferase‐like (QPCTL) is an isoenzyme of glutaminyl‐peptide cyclotransferase (QPCT). QPCTL and QPCT catalyze the formation of N‐terminal modified pyroglutamate‐fractalkine and the chemokine CCL2. The objective of this study was to investigate the association between insertions/deletions in the chicken QPCTL promoter region with growth traits in chickens. We first detected two insertion/deletion variants of QPCTL via whole‐genome resequencing analysis of DNA samples from Xichuan chickens. A total of 1896 individuals from 12 breeds were genotyped for 52‐ and 224‐bp insertions/deletions. We found two novel insertions/deletions in the promoter region of the chicken QPCTL gene and studied their association with chicken body weight and carcass traits. Our findings show that QPCTL can be a molecular marker for chicken genetics and breeding programs.     

A Single Nucleotide Polymorphism of Chicken Acetyl-CoA Carboxylase A Gene Associated with Fatness Traits

Acetyl-CoA carboxylase α (ACCα) is a major rate-limiting enzyme in the biogenesis of long-chain fatty acids. It can catalyze the carboxylation of acetyl-CoA to form malonyl-CoA that plays a key role in the regulation of fatty acid metabolism. The objective of the present study was to investigate the associations of ACCα gene polymorphisms with chicken growth and body composition traits. The Northeast Agricultural University broiler lines divergently selected for abdominal fat content and the Northeast Agricultural University F₂ Resource Population were used in the current study. Body weight and body composition traits were measured in the aforementioned two populations. A synonymous mutation was detected in the exon 19 region of ACCα gene, then polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method was developed to genotype all the individuals derived from the aforementioned populations. Association analysis revealed that the polymorphism was associated with abdominal fat weight and percentage of abdominal fat in the two populations. The results suggested that ACCα gene could be a candidate locus or linked to a major gene that affects abdominal fat content in the chicken.     

The Chicken Pan-Genome Reveals Gene Content Variation and a Promoter Region Deletion in IGF2BP1 Affecting Body Size

Domestication and breeding have reshaped the genomic architecture of chicken, but the retention and loss of genomic elements during these evolutionary processes remain unclear. We present the first chicken pan-genome constructed using 664 individuals, which identified an additional approximately 66.5-Mb sequences that are absent from the reference genome (GRCg6a). The constructed pan-genome encoded 20,491 predicated protein-coding genes, of which higher expression levels are observed in conserved genes relative to dispensable genes. Presence/absence variation (PAV) analyses demonstrated that gene PAV in chicken was shaped by selection, genetic drift, and hybridization. PAV-based genome-wide association studies identified numerous candidate mutations related to growth, carcass composition, meat quality, or physiological traits. Among them, a deletion in the promoter region of IGF2BP1 affecting chicken body size is reported, which is supported by functional studies and extra samples. This is the first time to report the causal variant of chicken body size quantitative trait locus located at chromosome 27 which was repeatedly reported. Therefore, the chicken pan-genome is a useful resource for biological discovery and breeding. It improves our understanding of chicken genome diversity and provides materials to unveil the evolution history of chicken domestication.     

Genetic Variants in ADD1 Gene and their Associations with Growth Traits in Cattle

The α-adducin (ADD1) is a subunit of adducin which is a cytoskeleton heterodimeric protein. Adducin participates in oocytes chromosome meiosis of mice, prompting adducin has an effect on embryonic development. Adducin gene mutation has significantly functional change. So the present study was to identify and characterize polymorphisms within the coding region of the bovine ADD1 gene among different cattle breeds. Here, 11 novel single nucleotide polymorphisms (SNPs 1–11) were identified by DNA sequencing and polymerase chain reaction-single stranded conformational polymorphism, there were one synonymous mutation in exon 1 (SNP1); four missense mutations in exons 4, 7, and 8 (SNPs 3–6); and six mutations in introns 4, 12, 13, and 14 (SNPs 2, 7–10). The statistical analyses indicated that the some SNPs are associated with the growth traits (body length, body height, chest circumference, and hucklebone width) in Chinese Jiaxian cattle population. Our results provide evidence that polymorphisms in the ADD1 gene are associated with growth traits, and may be used for marker-assisted selection in beef cattle breeding program.     

Correlation Analysis Between Single-Nucleotide Polymorphism of Malate Dehydrogenase Gene 5′-Flanking Region and Growth and Body Composition Traits in Chicken

Malate dehydrogenase (MD) is a key enzyme that plays an important role in energy metabolism. It catalyzes the oxidative decarboxylation of L-malate to yield CO₂ and pyruvate, while simultaneously generating NADPH from NADP⁺. The NADPH generated can be utilized in de novo synthesis of palmitate, which is the precursor molecule for the formation of other long-chain fatty acids. And high levels of MD will also activate muscle development. The current study was designed to investigate the effects of MD gene on growth and body-composition traits in chicken. The eighth generation population of Northeast Agricultural University broiler lines divergently selected for its abdominal fat and Northeast Agricultural University F₂ resource population were used in the research. Polymorphisms were detected by DNA sequencing and PCR-RFLP method was then developed to screen the population. A single mutation at the position of the 235 bp (Accession No. U49693) of MD 5′-flanking region was found. The correlation analysis between the polymorphism of the MD gene and growth and body composition traits was carried out using the appropriate statistic model. Least-square analysis showed that the BB genotype birds had much higher pectoralis major weight and percentage of pectoralis major than AA genotype birds (P<0.05). The abdominal fat weight, percentage of abdominal fat, the liver weight and percentage of liver weight of the AA genotype birds were much higher than those of BB genotype birds (P<0.05). These results indicate that MD gene is the major gene or is linked to the major gene that affects the growth and body composition traits in chicken.     

Gene structure and spatio-temporal expression of chicken LPIN2

LPIN2 is one of the members of the Lipin family, which acts as a phosphatidate phosphatase enzyme. In this study, we identified the cDNA sequence and exonic variants of chicken LPIN2, and evaluated its spatio-temporal expression patterns. It indicated that chicken LPIN2 cDNA contained a 2,664-bp open reading frame flanked by a 176-bp 5′ untranslated region and a 429-bp 3′ untranslated region, predicted encoding one protein of 886 amino acids. Fourteen variants (three missense mutations) were detected from the coding region of chicken LPIN2. W265L was predicted to affect the gene function (p < 0.01) and eight synonymous mutations were predicted to affect the binding sites of SR proteins, which suggested the important functions of these variants. Real-time quantitative PCR revealed that LPIN2 in two genotypic chickens (LD and HB chickens, with difference in growth rate) presented similar tissue expression patterns, which was liver and ovary enriched with low abundance in skeleton muscles. Chicken LPIN2 exhibited tissue-specific temporal-expression patterns during postnatal development (0–16 weeks). Chicken cutaneous LPIN2 was in steady-state mRNA levels during postnatal development; chicken LPIN2 mRNA in pectoralis major had a prominent level at 0 week-old, then dropped dramatically at 4 week-old and maintained a relatively low level through 4–16 weeks; while chicken hepatic LPIN2 had a relatively high expression at 0 week-old, with a relatively low level through 4–12 weeks and a slight increase at 16 week-old. The studies about the basic gene features of chicken LPIN2 would lay the foundation for further exploring its biological function.     

Quantitative trait loci segregating in crosses between New Hampshire and White Leghorn chicken lines: IV. Growth performance

Reciprocal crosses between the inbred lines New Hampshire (NHI) and White Leghorn (WL77) comprising 579 F₂ individuals were used to map QTL for body weight and composition. Here, we examine the growth performance until 20 weeks of age. Linkage analysis provided evidence for highly significant QTL on GGA1, 2, 4, 10 and 27 which had specific effects on early or late growth. The highest QTL effects, accounting for 4.6–25.6% of the phenotypic F₂ variance, were found on the distal region of GGA4 between 142 and 170 cM (F ≥ 13.68). The NHI QTL allele increased body mass by 141.86 g at 20 weeks. Using body weight as a covariate in the analysis of body composition traits provided evidence for genes in the GGA4 QTL region affecting fat mass independently of body mass. The QTL effect size differed between sexes and depended on the direction of cross. TBC1D1, CCKAR and PPARGC1A are functional candidate genes in the QTL peak region. Our study confirmed the importance of the distal GGA4 region for chicken growth performance. The strong effect of the GGA4 QTL makes fine mapping and gene discovery feasible.     

Molecular cloning and SNP association analysis of chicken PMCH gene

The pre-melanin-concentrating hormone (PMCH) gene is an important gene functionally concerning the regulations of body fat content, feeding behavior and energy balance. In this study, the full-length cDNA of chicken PMCH gene was amplified by SMART RACE method. The single nucleotide polymorphisms (SNPs) in the PMCH gene were screened by comparative sequence analysis. The obtained non-synonymous coding SNPs (ncSNPs) were designed for genotyping firstly. Its effects on growth, carcass characteristics and meat quality traits were investigated employing the F₂ resource population of Gushi chicken crossed with Anak broiler by AluI CRS-PCR–RFLP. Our results indicated that the cDNA of chicken PMCH shared 67.25 and 66.47 % homology with that of human and bovine PMCH, respectively. The deduced amino acid sequence of chicken PMCH (163 amino acids) were 52.07 and 50.89 % identical to those of human and bovine PMCH, respectively. The PMCH protein sequence is predicted to have several functional domains, including pro-MCH, CSP, IL7, XPGI and some low complexity sequence. It has 8 phosphorylation sites and no signal peptide sequence. gga-miR-18a, gga-miR-18b, gga-miR-499 microRNA targeting site was predicted in the 3′ untranslated region of chicken PMCH mRNA. In addition, a total of seven SNPs including an ncSNP and a synonymous coding SNP, were identified in the PMCH gene. The ncSNP c.81 A > T was found to be in moderate polymorphic state (polymorphic index = 0.365), and the frequencies for genotype AA, AB and BB were 0.3648, 0.4682 and 0.1670, respectively. Significant associations between the locus and shear force of breast and leg were observed. This polymorphic site may serve as a useful target for the marker assisted selection of the growth and meat quality traits in chicken.     

Polymorphisms of Flanking Region of the ASB15 Gene and Their Associations with Performance Traits in Chicken

Research on the identity of genes and their relationship with traits of economic importance in chickens could assist in the selection of poultry. In this study, an F₂ resource population of Gushi chickens crossed with Anka broilers was used to detect single-nucleotide polymorphisms (SNPs) in the flanking region of the ASB15 gene by DNA sequencing and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). One SNP of ?1271 C>T in 5′ flanking region of the chicken ASB15 gene and two SNPs of the 10618 A>G and 10716 G>A in 3′ flanking region were identified. Furthermore, the 10618 A>G and 10716 G>A in 3′ flanking region were in complete linkage. Association analysis results showed that ?1271 C>T was not associated with performance traits, while the 10618 A>G and 10716 G>A were significantly associated with BW2, 4, 6, 8, 10, 12, SL12, CD8, CW4, 8, 12, BSL4, 8, 12, and SEW, EW, WW, BMW, LW, CW, SFT. Our results suggest that the ASB15 gene profoundly affects chicken performance traits.     

Single-nucleotide polymorphisms in the promoter of the growth hormone-releasing hormone receptor gene are associated with growth and reproduction traits in chickens

Growth hormone‐releasing hormone receptor (GHRHR) plays a critical role in growth hormone (GH) synthesis, release and regulation in animals. The objective of this study was to investigate variations of the chicken GHRHR gene and their associations with growth and reproduction traits in 768 Beijing You chickens. Results revealed three single nucleotide polymorphisms (SNPs) in the promoter region of the gene (g.‐1654A>G, g.‐1411A>G and g.‐142T>C). Association analysis revealed that the novel SNP g.‐1654A>G had significant effects on chicken body weight at 7, 9, 11, 13, 17 weeks of age and the age of first egg as well as egg number at 32, 36 and 40 weeks. Significant association was also observed between g.‐1411A>G and g.‐142T>C with EN24. Moreover, the age of first egg was distinctly related with g.‐142T>C (P < 0.05). Although significant statistical difference was not detected in GHRHR mRNA levels among genotypes of the SNPs (P > 0.05), strong expression variations of the gene were found between the ages 17 and 20 weeks in the population (P < 0.05). These results suggest that the three SNPs in the GHRHR promoter could be used as potential genetic markers to improve the growth and reproductive traits in chickens.     

Polymorphisms of the Myostatin Gene and Its Relationship with Reproduction Traits in the Bian Chicken

Myostatin, or growth and differentiation factor 8, is a member of the transforming growth factor-β superfamily; it functions as a negative regulator of skeletal muscle development and growth in mammals. In this study, single nucleotide polymorphisms in the 5′ regulatory region and exon 1 of the myostatin gene were detected by PCR–SSCP in the Bian, Jinghai, Youxi, and Arbor Acre chickens, and the associations of the polymorphisms with reproduction traits were analyzed. Seven SNPs (A326G, C334G, C1346T, G1375A, A1473G, G1491A, and G2283A) were found in the myostatin gene. Association analysis showed that the G2283A were significantly associated with reproduction traits. Bian chickens of the GG genotype had a greater age at first egg than those of the GA and AA genotypes (P < 0.01). Correspondingly, Bian chickens of the GA and AA genotypes had larger egg number at 300 days than those of the GG genotype (P < 0.05 and P < 0.01, respectively). Bian chickens of the AA genotype had significantly higher body weight at 300 days than those of the GG genotype (P < 0.05). These results suggested that the myostatin gene may have certain effects on reproduction traits other than merely as a negative regulator of skeletal muscle development and growth in mammals previously reported.     

Changes in the Pattern of Twisted Gastrulation Gene Expression Among Drosophila Species

A long-standing hypothesis posits that morphological changes may be more likely to result from changes in regulation of gene expression than from changes in the protein coding sequences of genes. We have compared the expression pattern of the twisted gastrulation (tsg) gene among five Drosophila species: D. melanogaster, D. simulans, D. subobscura, D. mojavensis, and D. virilis. The tsg gene encodes a secreted protein that is required for the specification of dorsal midline fates in the Drosophila early embryo. TSG is unlike other secreted growth and differentiation factors in Drosophila in that its expression pattern can be experimentally varied and still result in normal development. Because of this, its regulatory region may be freer to diverge than that of other developmental genes whose misexpression may lead to lethal defects. Thus, the tsg gene may be a good indicator of the frequency and nature of evolutionary changes affecting patterns of gene expression. Over ∼60 million years (Myr), the tsg gene has retained a dorsal-on/ventral-off pattern and a middorsal region of expression; but there have been marked changes in the middorsal domain of expression as well as the appearance/loss of other domains of expression along the anterior/posterior axis. Changes between closely related species (∼2–5 Myr since divergence) that are not reflected among more distantly related species suggest frequent changes in gene expression over evolutionary time. These changes in gene expression may serve as the raw material for eventual evolutionary changes in morphology. Received: 24 March 1997 / Accepted: 20 June 1997