Isolation and molecular characterization of the stearoyl-CoA desaturase (SCD) gene affecting fat deposition in pigs

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Stearoyl-CoA desaturase (SCD) is a rate-limiting enzyme in the biosynthesis of unsaturated fatty acids. So far the porcine SCD gene has not been characterized. The isolation and molecular characterization of the full-length cDNA and genomic DNA sequence of the porcine SCD gene were described in the thesis. The 5134 bp cDNA contains a 1080 bp open reading frame encoding a protein of 359 amino acids with a calculated molecular mass of 41.3 kDa and theoretical isoelectric point of 9.4. The porcine SCD shares high identity (>80%) with the other mammalian SCD. To further elucidate the genomic structure of the porcine SCD gene, 20,985 bp of genomic DNA sequence was determined encompassing the complete porcine SCD gene.Similar to the other mammalian orthologs, particularly in term of exon size and exon/intron boundary, the porcine SCD gene spans a transcription unit of 16,186 bp, consisting of six exons with sizes ranging from 131 bp to 4048 bp, and five introns varying in size from 518 bp to 4784 bp. The unusual long 3" UTR of 3848 bp as opposed to the 176 bp 5' UTR appears in the last exon. A comparative analysis of different mammalian SCD promoters identified some regulatory domains required for the transcription regulation in the 5' flanking sequence of the porcine SCD gene, such as the conserved polyunsaturated fatty acid response region (PUFA-RE). Reverse transcription (RT)-PCR result indicates that the SCD gene is expressed ubiquitously in pigs. The porcine SCD gene was assigned to chromosome 14q27 by fluorescence in situ hybridization (FISH) and screening of hybrid panels with intronic primers.A total of 26 gene polymorphisms were revealed in the 21 kb DNA sequence, including 24 single nucleotide polymorphisms (SNPs), a 24 bp length polymorphism in the fourth intron and a triplet nucleotide insertion in the fifth intron. None of SNPs lead to an amino acid exchange. Significant differences in allele frequencies of the SNP T(-233)C and the SNP C(641)T (number refer to the corresponding position starting with +1 at the adenine of initiation codon ATG) were observed in samples of three Western commercial pig breeds (Landrace, Large White, and Duroc) and three Chinese indigenous pig breeds (Erhualian, Luchuan, and Huai).The Western pig breeds revealed higher frequencies of the allele T at the SNP T(-233)C, whereas the Chinese pig breeds showed higher frequencies of the allele T at the SNP C(641)T. Associations of SNPs T(-233)C and C(641)T with fatness traits have been investigated in F2 animals of a White Duroc × Erhualian pig resource family and a purebred White Duroc population. Significant associations were observed between the SNP C(641)T and backfat thickness at the 6-7th rib, backfat thickness at the last rib, and the average backfat thickness at four points (at the shoulder, the 6-7th rib, the last rib and the hip joint) in the F2 resource family (P<0.05). The allele T has an unfavorable effect (i.e. positive) on backfat thickness.