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The transcription factor csal1 is an important molecule that plays a critical regulatory function in ovarian follicle development, as confirmed by our previous data. However, the candidate genes of csal1 and its regulatory mechanism remain poorly understood in the granulosa cells (GCs) of chicken prehierarchical follicles (PFs). Six transcriptomes of csal1 and empty vector were analyzed in Chinese Dagu hens by RNA sequencing. Six cDNA libraries were constructed, with more than 42 million clean reads and 16,779 unigenes. Of these 16,779 unigenes, 2,762 differentially expressed genes (DEGs) were found in GCs, including 1,605 upregulated and 1,157 downregulated unigenes. Fourteen genes, including BMP5, TACR2, AMH, PLAG1, MYOD1, BOP1, SIPA1, NOTCH1, BCL2L1, SOX9, ADGRA2, WNT5A, SLC7A11, and GATAD2B, were related to GC proliferation and differentiation, hormone production, ovarian follicular development, regulation of reproductive processes, and signaling pathways in the PFs. Further analysis demonstrated the DEGs in GCs of ovarian follicles were enriched in neuroactive ligand-receptor interaction, cell adhesion molecules, and pathways related to cytochrome P450, indicating a critical function for csal1 in the generation of egg-laying features by controlling ovarian follicle development. For the first time, the current study represents the transcriptome analysis with ectopic csal1 expression. These findings provide significant evidence for investigating the molecular mechanism by which csal1 controls PF development in the hen ovary.

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The periostin (POSTN) and platelet-derived growth factor receptor-like (PDGFRL) genes are implicated in regulation of hen ovarian development. In the present study, these genes were explored as possible molecular markers associated with egg production, egg weight and body weight in Chinese Dagu hens. Samples were analyzed using the PCR-single strand conformation polymorphism (PCR-SSCP) method, followed by sequencing analysis, and three novel single nucleotide polymorphisms (SNPs) were identified within the candidate genes. Among them, an A/T transversion at base position 2727 in intron 2 of POSTN gene was found to be polymorphic and named SNP A2727T; and two transitions, G/A at position 6761 and A/G at base 6839 in exon 2 of PDGFRL gene were detected and named SNPs G6761A and A6839G, respectively. For the SNP A2727T, a total of 360 Dagu hens were classified as AA and AB genotypes, allele A was found present at a higher frequency. Moreover, the AA genotype was significantly correlated with higher hen-housed egg production (HHEP) at 43, 57, and 66 weeks (wks) of age and with a higher egg weight (EW) at 30 wks (P < 0.05). For the two linked SNPs (G6761A and A6839G) in the PDGFRL fragment, the hens were typed into TT, TC and CC genotypes, with the T allele shown to be dominant. The TT genotype was correlated with higher HHEP at 57 and 66 wks of age; genotype CC associated with the highest body weight and EW at 30 and 43 wks (P < 0.05), while it was correlated with the lowest HHEP at 57 and 66 wks of age (P < 0.05). Furthermore, five haplotypes were reconstructed based on these SNPs, with the AATT haplotype associated with the highest HHEP at 43 to 66 wks of age and higher EW at 30 wks (P < 0.05). Collectively, these SNPs identified in this study might be used as a potential molecular marker favorable to genetic improvement of egg productivity in chicken breeding.

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Adrenergic, alpha-1B-, receptor (ADRA1B) and peroxisome proliferator-activated receptor gamma, coactivator 1 beta (PPARGC1B) genes are involved in regulation of hen ovarian development. In this study, these two genes were investigated as possible molecular markers associated with hen-housed egg production, egg weight (EW) and body weight in Chinese Dagu hens. Samples were analyzed using the polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) technique, followed by sequencing analysis. Two novel single nucleotide polymorphisms (SNPs) were identified within the candidate genes. Among them, an A/G transition at base position 1915 in exon 2 of ADRA1B gene and a T/C mutation at base position 6146 in the 3'-untranslated region (UTR) of PPARGC1B gene were found to be polymorphic and named SNP A1915G and T6146C, respectively. The SNP A1915G (ADRA1B) leads to a non-synonymous substitution (aspartic acid 489-to-glycine). The 360 birds from the Dagu population were divided into genotypes AA and AG, allele A was found to be present at a higher frequency. Furthermore, the AG genotype correlated with significantly higher hen-housed egg production (HHEP) at 30, 43, 57, and 66 wks of age and with a higher EW at 30 and 43 wks (p<0.05). For the SNP T6146C (PPARGC1B), the hens were typed into TT and TC genotypes, with the T allele shown to be dominant. The TC genotype was also markedly correlated with higher HHEP at 57 and 66 wks of age and EW at 30 and 43 wks (p<0.05). Moreover, four haplotypes were reconstructed based on these two SNPs, with the AGTC haplotype found to be associated with the highest HHEP at 30 to 66 wks of age and with higher EW at 30 and 43 wks (p<0.05). Collectively, the two SNPs identified in this study might be used as potential genetic molecular markers favorable in the improvement of egg productivity in chicken breeding.

RESUMEN

Transcription factor forkhead box L2 (FOXL2) and growth differentiation factor-9 (GDF9) genes have critical roles in the regulation of hen ovarian development. In the present study, these genes were explored as possible molecular markers associated with BW, hen-housed egg production, and egg weight in Chinese Dagu hens. Samples were analyzed using the PCR-single strand conformation polymorphism (PCR-SSCP) technique followed by sequencing analysis, and two novel single nucleotide polymorphisms (SNPs) were identified within these candidate genes. Among them, an A/G transition at base position 238 in the coding region of the FOXL2 gene and a G/T transversion at base position 1609 in exon 2 of the GDF9 gene were found to be polymorphic and named SNPs A238G and G1609T, respectively. The SNP A238G (FOXL2) leads to a nonsynonymous substitution (isoleucine77-to-valine), and when the 360 Dagu hen samples were divided into genotypes AA and AB, allele A was found to be present at a higher frequency. Furthermore, the AA genotype correlated with significantly higher hen-housed egg production at 30, 43, 57, and 66 wk of age and with a higher egg weight at 43 wk (P<0.05). For the SNP G1609T (GDF9), the hens were typed into TT and TC genotypes, with the T allele shown to be dominant. The TC genotype was also markedly correlated with higher hen-housed egg production and a higher egg weight (P<0.05). Moreover, four haplotypes were reconstructed based on these two SNPs, with the AATC haplotype found to be correlated with the highest hen-housed egg production at 30 to 66 wk of age and with higher egg weights at 43 wk (P<0.05). Collectively, the two SNPs identified in this study might be used as possible genetic molecular markers to aid in the improvement of egg production traits in chicken breeding.

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