Novel single nucleotide polymorphisms of insulin-like growth factor-binding protein 7 (IGFBP7) gene significantly associated with growth traits in striped catfish (Pangasianodon hypophthalmus Sauvage, 1878).

Breeding program to improve economically important growth traits in striped catfish (Pangasianodon hypophthalmus) requires effective molecular markers. This study was conducted to identify single nucleotide polymorphisms (SNPs) of Insulin-like Growth Factor-Binding Protein 7 (IGFBP7) gene which plays multiple roles in regulating growth, energy metabolism and development. The association between SNPs in IGFBP7 gene and growth traits in striped catfish was analyzed in order to uncover the SNPs that have potential to be valuable markers for improving growth traits. Firstly, fragments of IGFBP7 gene from ten fast-growing fish and ten slow-growing fish were sequenced in order to discover SNPs. After filtering the detected SNPs, an intronic SNP (2060A > G) and two non-synonymous SNPs (344 T > C and 4559C > A) causing Leu78Pro and Leu189Met in protein, respectively, were subjected to further validated by individual genotyping in 70 fast-growing fish and 70 slow-growing fish using single base extension method. Our results showed that two SNPs (2060A > G and 4559 C > A (p. Leu189Met)) were significantly associated with the growth in P. hypophthalmus (p < 0.001), thus being candidate SNP markers for the growth traits of this fish. Moreover, linkage disequilibrium and association analysis with growth traits of haplotypes generated from the 3 filtered SNPs (344 T > C, 2060 A > G and 4559 C > A) were examined. These revealed that the non-coding SNP locus (2060A > G) had higher genetic diversity at which the G allele was predominant over the A allele in the fast-growing fish. Furthermore, the results of qPCR showed that expression of IGFBP7 gene with genotype GG (at locus 2060) in fast-growing group was significantly higher than that with genotype AA in slow-growing group (p < 0.05). Our study provides insights into the genetic variants of IGFBP7 gene and useful data source for development molecular marker for growth traits in breeding of the striped catfish.

The complete mitochondrial genome sequence of the indigenous I pig (Sus scrofa) in Vietnam.

OBJECTIVE: The I pig is a long nurtured longstanding breed in Vietnam, and contains excellent indigenous genetic resources. However, after 1970s, I pig breeds have become a small population because of decreasing farming areas and increasing pressure from foreign breeds with a high growth rate. Thus, there is now the risk of the disappearance of the I pigs breed. The aim of this study was to focus on classifying and identifying the I pig genetic origin and supplying molecular makers for conservation activities. METHODS: This study sequenced the complete mitochondrial genome and used the sequencing result to analyze the phylogenetic relationship of I pig with Asian and European domestic pigs and wild boars. The full sequence was annotated and predicted the secondary tRNA. RESULTS: The total length of I pig mitochondrial genome (accession number KX094894) was 16,731 base pairs, comprised two rRNA (12S and 16S), 22 tRNA and 13 mRNA genes. The annotation structures were not different from other pig breeds. Some component indexes as AT content, GC, and AT skew were counted, in which AT content (60.09%) was smaller than other pigs. We built the phylogenetic trees from full sequence and D loop sequence using Bayesian method. The result showed that I pig, Banna mini, wild boar (WB) Vietnam and WB Hainan or WB Korea, WB Japan were a cluster. They were a group within the Asian clade distinct from Chinese pigs and other Asian breeds in both phylogenetic trees (0.0004 and 0.0057, respectively). CONCLUSION: These results were similar to previous phylogenic study in Vietnamese pig and showed the genetic distinctness of I pig with other Asian domestic pigs.

A single amino acid substitution alters omnipotent eRF1 of Dileptus to euplotes-type dualpotent eRF1: standard codon usage may be advantageous in raptorial ciliates.

Most ciliates use a deviant genetic code. Eukaryotic release factor (eRF1) appears to play an important role in the process of reassignment of stop codons. Although the precise site on eRF1 for recognition of stop codons remains obscure, studies have suggested that the tip region NIKS and its adjacent YxCxxxF motifs in domain 1 are important for stop codon recognition. Litostomatea is a class of ciliate that appears to use the standard genetic code. We used Dileptus (Litostomatea) eRF1 in this study to identify key residues located in or near the YxCxxxF motif. We predicted sites involving stop codon recognition by computational calculation of RNA-protein interaction propensity. We introduced mutations at the predicted sites of Dileptus eRF1 and examined the activity of the mutated Dileptus eRF1 using in vivo assay systems. The results show that the single mutation R128I (Dileptus eRF1 numbering) in the YxCxxxF motif converted the omnipotent recognition of Dileptus eRF1 to Euplotes-type dualpotent eRF1. Our results indicate that R128 is one of the key residues preserving the ability to recognize all three stop codons, especially UGA, in Dileptus. We discuss a possible advantage that ciliates from the Litostomatea class may gain from using the standard genetic code.

Ciliates use both variant and universal genetic codes: evidence of omnipotent eRF1s in the class Litostomatea.

Stop codon reassignments have occurred very frequently in ciliates. In some ciliate species, the universal stop codons UAA and UAG are translated into glutamine, while in some other species, the universal stop codon UGA appears to be translated into cysteine or tryptophan. The class Litostomatea has been hypothesized to be the only group of ciliates using the universal genetic code. However, the hypothesis was based on a statistical analysis of quite small sequence dataset which was insufficient to elucidate the codon usage of the class among such highly deviated phylum. In this study, together with the updated database sequence analysis for the class, we approached the problem of stop codon usage by examining the capacity of the translation termination factor eRF1 for recognizing stop codons. Using in vivo assay systems in budding yeast, we estimated the activity of eRF1 from two litostome species Didinium nasutum and Dileptus margaritifer. The results clearly showed that Didinium and Dileptus eRF1s efficiently recognize all three stop codons. This is the first experimental evidence that strongly supports the hypothesis that litostome ciliates use universal genetic code.

Newly sequenced eRF1s from ciliates: the diversity of stop codon usage and the molecular surfaces that are important for stop codon interactions.

The genetic code of nuclear genes in some ciliates was found to differ from that of other organisms in the assignment of UGA, UAG, and UAA codons, which are normally assigned as stop codons. In some ciliate species, the universal stop codons UAA and UAG instead encode glutamine. In some other ciliates, the universal stop codon UGA appears to be translated as cysteine or tryptophan. Eukaryotic release factor 1 (eRF1) is a key protein in stop codon recognition, thus, the protein is believed to play an important role in the stop codon reassignment in ciliates. We have cloned, sequenced, and analyzed the cDNA of eRF1 from four ciliate species of three different classes: Karyorelictea (Loxodes striatus), Heterotrichea (Blepharisma musculus), and Litostomatea (Didinium nasutum, Dileptus margaritifer). Phylogenetic analysis of these eRF1s supports the hypothesis that the genetic code in ciliates has deviated independently several times from the universal genetic code, and that different ciliate eRF1s may have undergone different processes to change the codon specificity. Using computational methods, we have also suggested areas on the surface of eRF1s that are important for stop codon recognition in ciliate eRF1s.