| Crested ibis considered as one of the four national treasures in China, is a kind of critically endangered bird. It was almost extinctive during 1960's ? 1970's until a population of crested ibis consisted of seven birds was found in Yangxian county of Shaanxi province. Up to now, there are four populations in Yangxian, Zhouzhi county of Shaanxi, Beijing zoo in China and Japan, and the total number is up to one thousand. Although it is a successful example in word in endangered animal protection using modern management and technologies, it is necessary to improved and amplify the population of crested ibis. In this study, Genetic variations of FSH, TSH of glycoprotein, BDNF, GDNF genes of NT family, 12S rRNA and c-mos genes in crested ibis came from Yangxian and Zhouzhi county of Shaanxi province were detected using PCR-SSCP DNA marker technique. The objective of this study was to invest the genetic characteristics of the crested ibis population, to provide information for improvement and amplification of crested ibis and the studies on reproduction, development and mechanisms of some diseases of crested ibis, so the protection of crested ibis would be carried out scientifically. The results were as follows:1. BDNF gene of 63 crested ibis was analyzed and one silent mutation (G→A,coding Ala) was found in the site 450. There were two genotype AA and AB were found and the frequency were 0.7143 and 0.2857, respectively. Allele frequency of A/B were 0.8571/0.1429, respectively. Chi-square test showed that this locus was in Hardy?Weinberg equilibrium. The genetic diversity indexes of H/PIC were 0.2450/0.2149 in the crested ibis population. The genetic variance of this locus was low, which indicated lower genetic variance information in these crested ibis. GDNF gene of 63 crested ibis was detected by PCR-SSCP and only 2?band haplotype was found. No polymorphism was found. Homology analyses were carried out among the BDNF sequences of crested ibis and other species. 405 mutation loci and 342 degeneracy loci were found among 820 loci and 49.4 percent and 41.7 percent, respectively. The transition was more than transversion among species sequence variant. The phyletic tree of crested ibis, amphibian and teleostei showed that crested ibis belong to archosauria of Euteleostomi and nearer to lepidosauria and farer to mammalian.2. Phylogenesis of NTs family BDNF,GDNF,NT3 and NT5 genes of 39 species was carried out, which the phyletic relationship was (GDNF,(NT5,(NT3,BDNF))). The lower homology of this family indicated that every gene had a rapid change speed.3. A SNP (C→T) was detected in FSHβ5′flanking region. Only genotype AA and AB were found in the population. Genotypic frequencies of AA and AB were 0.7619 and 0.2381, respectively. Allele frequencies of A/B were 0.8810 and 0.1190, respectively. Genotypic AA and allele A were dominant in this population. The chi-square test showed the population was at Hardy-Weinberg disequilibration (P<0.05). The genetic diversity indexes of H/PIC were 0.2097/7.7561 in the crested ibis population. So the population was low polymorphic at this SNP locus. No polymorphism was detected in exon 2 in FSHβsubunit in crested ibis population. The fragment of FSHβexon 2 was sequenced and phylogenetic analysis results showed that the high sequence similarities of exon 2 in FSHβgene of crested ibis with mallard, chicken and quail were 92.09%, 91.37% and 89.93%, respectively. Phylogenetic analysis indicated crested ibis was cluster to mallard and far from quail.4. A SNP (C→T) was detected in 5′flanking region of TSHβgene. There were three genotypes were found in the population and genotype AA, AB and BB frequencies were 0.3651, 0.5714 and 0.0635, respectively. Allele frequencies of A/B were 0.6508/0.3492. The chi-square test showed this population was at Hardy-Weinberg disequilibration (P<0.05). The genetic diversity indexes of H/PIC were 0.4545/0.3512 in the crested ibis population. So the locus was moderate polymorphic. A SNP (T→A) was detected in exon 2 of TSHβgene and two genotypes were found in the population. Genotype AA and AB frequencies were 0.4127, 0.5873, respectively. Allele frequencies of A/B were 0.7063/0.2937. The chi-square test showed this population was at Hardy-Weinberg critically disequilibration (P<0.01). The allele B faced to the disappear tendency. The genetic diversity indexes of H/PIC were 0.4148/0.3288 in the crested ibis population. So the locus was moderate polymorphic. The two loci indicated the genetic variation of crested ibis recovered to a certain extent.5. Phyletic tree of vertebrate glycoprotein hormonesβsubunit nucleotide sequences among 51 species wasconstructed. The results showed that hCG and LH had nearer relationship, and gradually farer with FSH and TSH. While the phyletic tree of nucleotide sequences showed that the difference was larger between these two groups than TSH and FSH, hCG and LH. The results indicated FSH, TSH, LH and hCG had higher homology and lower phyletic evolution speed.6. c-mos gene partial sequence of 63 crested ibis was analyzed and one silent mutation (C→T,coding Tyr) was found in the site 282. There were three genotype AA, AB and BB were found and the frequency were 0.5873, 0.2857 and 0.1270, respectively. Allele frequencies of A/B were 0.7302/0.2698, respectively. Chi-square test showed that this locus was in Hardy?Weinberg disequilibrium(P<0.05). The genetic diversity indexes of H/PIC were 0.3940/0.3164 in the crested ibis population, which indicated this locus was a midrange locus. Phylogenetic analysis of ciconiiformes based on analysis of the nuclear c-mos gene, mitochondrial 12S rRNA gene sequences and the combined sequences of the tow genes. The result showed that combined sequences analysis was more reliable than single sequence analysis. The Phylogenetic of ciconiiformes was (Threskiornithidae((Balaenicipitidae,Scopidae)Cathartidae, Ciconiidae, Ardeidae)). No polymorphism was detected in three loci of 12S rRNA sequence.7. Five polymorphic loci of 6 genes were comprehensive analyzed and the results showed that H|- and PIC|─were 0.3436 and 0.2798, respectively, which indicated that the two groups of Yangxian and Zhouzhi county of Shaanxi province still had a lower genetic variant. We should work hard to protect this critically endangered bird, and ensure this specy recovery quickly. |
