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Biochemical Genetics (v.47, #1-2)


A Phylogroup of the Siberian Chipmunk from Korea (Tamias sibiricus barberi) Revealed from the Mitochondrial DNA Cytochrome b Gene by Hung Sun Koh; Jinxing Wang; Bae Kun Lee; Byong Guk Yang; Seon Wook Heo; Kyung Hee Jang; Tae Yong Chun (pp. 1-7).
In the full sequences of the mtDNA cytochrome b gene, 26 haplotypes (Tamias sibiricus barberi) from six localities of central and southern Korea were distinct from 21 haplotypes (Tamias sibiricus orientalis) from five localities of northeast China and Vladivostok, Russia. The average Tamura–Nei nucleotide distance between the subspecies (11.40%) and maximum infrasubspecific distances (3.74% and 4.72%) support the subspecies classification of T. s. barberi based on morphometric comparison. The 26 haplotypes of T. s. barberi were also distinct from 2 haplotypes of T. s. orientalis and Tamias sibiricus jacutensis from far-east Russia (average distance, 11.86%). Thus T. s. barberi constitutes a “phylogroup” (average nucleotide distance > 10%); analyses with nuclear genes of northeast Asian specimens, including North Korean ones, are necessary to clarify its taxonomic status. Furthermore, 49 haplotypes of T. sibiricus from eastern Asia differed from 19 haplotypes of another 18 Tamias spp. from America (weighted-average distance, 18.58%). T. sibiricus is, therefore, distinct enough to be recognized as a subgenus, Eutamias.

Keywords: Tamias sibiricus barberi ; Siberian chipmunk from Korea; Phylogroup; Cytochrome b gene


Relationships Among Genetic Makeup, Active Ingredient Content, and Place of Origin of the Medicinal Plant Gastrodia Tuber by Jun Tao; Zhi-yong Luo; Chikira Ismail Msangi; Xiao-shun Shu; Li Wen; Shui-ping Liu; Chang-quan Zhou; Rui-xin Liu; Wei-xin Hu (pp. 8-18).
Gastrodia tuber and its component gastrodin have many pharmacological effects. The chemical fingerprints and gastrodin contents of eight Gastrodia populations were determined, and the genomic DNA polymorphism of the populations was investigated. Genetic distance coefficients among the populations were calculated using the DNA polymorphism data. A dendrogram of the genetic similarities between the populations was constructed using the genetic distance coefficients. The results indicated that the genomic DNA of Gastrodia tubers was highly polymorphic; the eight populations clustered into three major groups, and the gastrodin content varied greatly among these groups. There were obvious correlations among genetic makeup, gastrodin content, and place of origin. The ecological environments in Guizhou and Shanxi may be conducive to evolution and to gastrodin biosynthesis, and more suitable for cultivation of Gastrodia tubers. These findings may provide a scientific basis for overall genetic resource management and for the selection of locations for cultivating Gastrodia tubers.

Keywords: DNA polymorphism; Gastrodia elata Blume; Gastrodin; Genetic relationships; Chemical fingerprints

Polymorphic Study of XRCC1 G28152A and XRCC1 C26304T in 10 Chinese Populations by Chao Wang; Feng Chen; Xuelong Zhang; Yan Jin; Jing Bai; Songbin Fu (pp. 27-32).

Analysis of the Alpha-1-Antitrypsin Deficient Alleles M3S, MZ, and ZZ by Biochemical and Molecular Methods: A Family Study by Mustafa Calapoğlu; Orhan Değer; Fulya Balaban; Nilufer Şahin Calapoğlu; Yılmaz Bülbül; Keith Burling (pp. 33-41).
Deficiency of alpha-1-antitrypsin (α1-AT, a major protease inhibitor controlling tissue degradation) is a genetic disorder transmitted in a codominant autosomal form. It has more than 100 genetically determined variants. This study attempted to determine the degree of association between serum α1-AT levels and phenotypes and to provide a strategy for reliable laboratory evaluation of deficiencies. The study group consisted of a 38-year-old male proband with clinical features of emphysema, his first-degree relatives, and healthy controls. Family history revealed a four-generation pedigree. Genomic DNA was isolated from peripheral blood leukocytes. Alpha-1-AT levels were determined from human serum by immunonephelometry. Phenotypes were determined by isoelectric focusing of blood samples. DNA sequences of coding exons were analyzed by the amplification DNA technique and direct sequencing. Inheritance and plasma levels of the ZZ, MM, M3S, and MZ phenotypes were confirmed by the family study. In the family members with deficiencies, plasma concentrations were 22.55% ± 5.15 (ZZ), 84.18% ± 5.18 (M3S), and 61.06% ± 7.15 (MZ) of the normal MM level. We found a close association between α1-AT level and genotype. A combination of genotyping, quantification, and phenotyping is the optimal strategy for the laboratory evaluation of α1-AT deficiency.

Keywords: Alpha-1-antitrypsin deficiency; Genetic screening; Phenotype; PI*M3 allele; Confidence intervals


Genetic Diversity and Geographic Differentiation of the Giant Tiger Shrimp (Penaeus monodon) in Thailand Analyzed by Mitochondrial COI Sequences by Bavornlak Khamnamtong; Sirawut Klinbunga; Piamsak Menasveta (pp. 42-55).
Genetic diversity and geographic differentiation of the giant tiger shrimp, Penaeus monodon, in Thai waters (Satun, Trang, Phangnga, and Ranong in the Andaman Sea and Chumphon and Trat in the Gulf of Thailand) were examined by COI polymorphism (N = 128). We observed 28 COI mitotypes across all investigated individuals. The sequence divergence between pairs of mitotypes was 0.00–20.76%. A neighbor-joining tree clearly indicated lineage separation of Thai P. monodon and large nucleotide divergence between interlineage mitotypes but limited divergence between intralineage mitotypes. High genetic diversity was found (mean sequence divergence = 6.604%, haplotype diversity = 0.716–0.927, π = 2.936–8.532%). F-statistics (F ST) and an analysis of molecular variance (AMOVA) indicated that the gene pool of Thai P. monodon was not homogeneous but genetically differentiated intraspecifically (P < 0.05). Six samples of P. monodon could be allocated into three different genetic populations: Trat (A), Chumphon (B), and the Andaman samples Satun, Trang, Phangnga, and Ranong (C). Contradictory results regarding patterns of geographic differentiation previously reported by various molecular approaches were clarified by this study.

Keywords: Genetic diversity; Geographic differentiation; Stock identification; Penaeus monodon ; COI


High ISSR Variation in 14 Surviving Individuals of Euryodendron excelsum (Ternstroemiaceae) Endemic to China by Yingjuan Su; Ting Wang; Yufei Sun; Huagu Ye (pp. 56-65).
Euryodendron excelsum is a critically endangered Ternstroemiaceae species endemic to southern China, with only 14 individuals surviving in Ba Jia Zhen of Yangchun, Guangdong Province. Intersimple sequence repeat (ISSR) markers were used to assess genetic variation and relationships among these individuals. Population genetic parameters were estimated by a Bayesian approach as well as conventional methods. Of the 225 loci generated by 21 primers, 147 (65.33%) were polymorphic. Compared with other species of Theaceae and related families, a high level of genetic variation was identified in E. excelsum (Nei’s gene diversity, 0.2458; Shannon’s index, 0.3626). An unweighted pair group method with arithmetic mean (UPGMA) dendrogram showed that the 14 individuals were mainly clustered into three groups, a conclusion further supported by principal coordinate analysis. Based on these results, a management and conservation strategy for E. excelsum was proposed.

Keywords: Euryodendron excelsum ; Genetic diversity; Conservation management


Isolation and Characterization of Testis-Specific DMRT1 in the Tropical Abalone (Haliotis asinina) by Sirawut Klinbunga; Piti Amparyup; Bavornlak Khamnamtong; Ikuo Hirono; Takashi Aoki; Padermsak Jarayabhand (pp. 66-79).
The Doublesex Male abnormal-3 Related Transcription factor-1 (DMRT1) gene encodes a protein containing the DNA-binding motif called the DM domain, involved in the sexual development of various species. To gain insight into its implications for gonadal differentiation in the tropical abalone (Haliotis asinina), a DMRT1 homolog was identified and characterized. The full length cDNA of HADMRT1 (1,740 bp with an ORF of 732 bp corresponding to a putative polypeptide of 243 amino acids) and its DM domain-less variant (HADMRT1-like, 1,430 bp with an ORF of 312 bp, 103 amino acids) were successfully isolated and reported for the first time in molluscs. HADMRT1 was specifically expressed in the testes of adult H. asinina (N = 16) but not in whole juveniles (2, 3, 5 months old, N = 6 for each group) and ovaries (N = 16), and pooled hemocytes (from 50 individuals) of adults. Tissue distribution analysis further revealed testis-specific expression of HADMRT1. Semiquantitative RT-PCR illustrated that the relative expression level of HADMRT1 in developed testes (stages II, III, and IV) was significantly greater than that in undeveloped testes (stage I) of abalone broodstock (P < 0.05).

Keywords: RACE-PCR; Testis-specific expression; DMRT1; DM domain-less; Abalone


On the Diversity of the Laccase Gene: A Phylogenetic Perspective from Botryosphaeria rhodina (Ascomycota: Fungi) and Other Related Taxa by Flávio J. D. Castilho; Rodrigo A. Torres; Aneli M. Barbosa; Robert F. H. Dekker; José E. Garcia (pp. 80-91).
The present study is the first describing the sequencing of a fragment of the copper-oxidase domain of a laccase gene in the family Botryosphaeriaceae. The aim of this work was to assess the degree of genetic and evolutionary relationships of a laccase gene from Botryosphaeria rhodina MAMB-05 with other ascomycete and basidiomycete laccase genes. The 193-amino acid sequences of the copper-oxidase domain from several different fungi, insects, a plant, and a bacterial species were retrieved from GenBank and aligned. Phylogenetic analyses were performed using neighbor-joining, maximum parsimony, and Bayesian inference methods. The organisms studied clustered into five gene clades: fungi (ascomycetes and basidiomycetes), insects, plants, and bacteria. Also, the topologies showed that fungal laccases of the ascomycetes and basidiomycetes are clearly separated into two distinct clusters. This evidence indicated that B. rhodina MAMB-05 and other closely related ascomycetes are a new biological resource given the biotechnological potential of their laccase genes.

Keywords: Botryosphaeria rhodina MAMB-05; Laccase; Ascomycete; Basidiomycete; Phylogenetic analyses; Copper-oxidase domain


Hox Genes in the Parasitic Platyhelminthes Mesocestoides corti, Echinococcus multilocularis, and Schistosoma mansoni: Evidence for a Reduced Hox Complement by Uriel Koziol; Ana I. Lalanne; Estela Castillo (pp. 100-116).
Little is known about the Hox gene complement in parasitic platyhelminthes (Neodermata). With the aim of identifying Hox genes in this group we performed two independent strategies: we performed a PCR survey with degenerate primers directed to the Hox homeobox in the cestode Mesocestoides corti, and we searched genomic assemblies of Echinococcus multilocularis and Schistosoma mansoni. We identified two Hox genes in M. corti, seven in E. multilocularis, and nine in S. mansoni (including five previously reported). The affinities of these sequences, and other previously reported Hox sequences from flatworms, were determined according to phylogenetic analysis, presence of characteristic parapeptide sequences, and unusual intron positions. Our results suggest that the last common ancestor of triclads and neodermatans had a Hox gene complement of at least seven genes, and that this was probably derived by gene loss from a larger ancestral Hox complement in lophotrochozoans.

Keywords: Hox ; Parahox; Platyhelminthes; Cestode; Mesocestoides corti ; Neodermata


Analysis of Expressed Sequence Tags in Porcine Uterus Tissue by Hui Chai; Dong-Liang Yu; Bing Zhang; Yan Fu; Song-Nian Hu (pp. 117-125).
Two non-normalized cDNA libraries of uteri from Danish Landrace and Chinese Erhualian pigs were constructed, and 13,756 expressed sequence tags (ESTs) were randomly sequenced. The ESTs were clustered by Phrap software, and 6,139 distinct tentative consensus sequences were produced, including 2,730 contigs and 3,409 singlets. Using Blast tools, these 6,139 candidate genes were compared to the nr and nt databases; 5,210 of them were assigned putative functions, whereas 929 potentially represent new genes. Highly expressed genes appear to be associated with basic energy metabolism, transferase activity, localization, cellular physiological process, protein binding, and nucleic acid binding. Antileukoproteinase was the most highly expressed gene, corresponding to endometrial differentiation and conceptus or fetal development.

Keywords: Porcine uterus; Porcine ESTs; Gene expression; Pig reproduction


Utilization of the Porcine System to Study Lymphotoxin-β Regulation in Intestinal Lymphoid Tissue by Tomoyuki Shimazu; Masanori Tohno; Shinichiro Katoh; Takeshi Shimosato; Hisashi Aso; Yasushi Kawai; Tadao Saito; Haruki Kitazawa (pp. 126-136).
Lymphotoxin-β (LT-β) has been suggested to be a regulator of secondary lymphoid structure development. In the present study, we isolated porcine LT-β (poLT-β) from adult swine spleens. The open reading frame encoded a predicted 246-amino acid polypeptide exhibiting higher similarity to the human than the mouse LT-β protein. Expression of LT-β mRNA in various swine tissues was analyzed by real-time PCR, and it was found to be higher in the ileal Peyer's patches (Pps) of adults than in newborns. In addition, ligand stimulation of toll-like receptors 2, 4, and 9, which are activated by bacterial components, increased LT-β expression only in neonatal ileal Pps. These results suggest that colonization by commensal bacteria may affect the maturation of neonatal ileal Pps by the induction of LT-β via toll-like receptors. LT-β may therefore be useful for studying the development of the intestinal immune system at parturition in both swine and humans.

Keywords: LT-β porcine; Toll-like receptors; GALT; Neonate


Isolation and Expression of a Novel Alligator Gene Belonging to the Sox Gene Family by Jifang Zheng; Nan Hu; Muyuan Zhu; Yaqing Nu; Zhen Liu (pp. 137-146).
Sox genes share a highly conserved DNA-binding motif, the HMG (high mobility group)-box domain, and have diverse roles in vertebrate embryonic development. A novel SRY-related cDNA (temporarily called Sox33) isolated from the Chinese alligator (Alligator sinensis) is 1,819 bp in length, with an open reading frame from 220 to 1113 bp, encoding a protein of 298 amino acids. Two putative polyadenylation signal sequences (AATAAA) are present upstream of the poly(A) tail in the 3′ UTR (at 1255–1260 and 1774–1779). The putative protein contains an HMG-box domain most closely related to hSox12, mSox4, rtSox11, and mSox11 homologs, indicating that alligator Sox33 belongs to group C in the Sox gene family. Alligator adult and developing tissues were tested for Sox33 mRNA by independent Northern blots using a 336-bp probe (at 907–1243) between the HMG-box and the poly(A) site I and a 277-bp probe (at 1477–1754) between the two polyadenylation sites. Two transcripts (1.3 kb and 1.8 kb) in developing brain and one (1.8 kb) in adult brain were identified by the 336-bp probe; only one transcript (1.8 kb) in developing and adult brains was detected by the 277-bp probe. The results suggest that alligator Sox33 may use a different polyadenylation mechanism in the developing brain and play a role in the development and maintenance of the nervous system.

Keywords: Alligator (Alligator sinensis); SRY ; Sox gene; Cloning; Expression

Sequence Analysis of Human TRPV6 Suggests Positive Selection Outside Africa by Mikiko Soejima; Hidenori Tachida; Yoshiro Koda (pp. 147-153).

Genetic Variation in Chinese Pine (Pinus tabulaeformis), a Woody Species Endemic to China by Meng-Ben Wang; Feng-Qin Gao (pp. 154-164).
Random amplified polymorphic DNA markers were used to investigate genetic variation of the Chinese pine (Pinus tabulaeformis Carr.), a species endemic to China and the most widely distributed pine species in North China. The results revealed that P. tabulaeformis populations had a relatively high level of genetic diversity (H t = 0.3268), distributed mainly within (79.2%) rather than among (20.8%) populations. The populations of Lingkong Mountain and Wuling Mountain had a higher level of diversity (0.2687) than the other four populations (0.2537). No statistically significant relationships were found between genetic diversity and climatic factors by correlation analysis and between genetic distance and geographic distance by the Mantel test. These results suggest that the partitioning of genetic diversity in each population might have been influenced not only by water and temperature conditions but also by other factors such as human activities and the Holocene postglacial history of these populations.

Keywords: RAPD; Pinus tabulaeformis ; Genetic diversity; Conservation; China
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