Purification and characterization of a chloride ion-dependent α-glucosidase from the midgut gland of Japanese scallop (Patinopecten yessoensis)

Marine glycoside hydrolases hold enormous potential due to their habitat-related characteristics such as salt tolerance, barophilicity, and cold tolerance. We purified an α-glucosidase (PYG) from the midgut gland of the Japanese scallop (Patinopecten yessoensis) and found that this enzyme has unique characteristics. The use of acarbose affinity chromatography during the purification was particularly effective, increasing the specific activity 570-fold. PYG is an interesting chloride ion-dependent enzyme. Chloride ion causes distinctive changes in its enzymatic properties, increasing its hydrolysis rate, changing the pH profile of its enzyme activity, shifting the range of its pH stability to the alkaline region, and raising its optimal temperature from 37 to 55 °C. Furthermore, chloride ion altered PYG’s substrate specificity. PYG exhibited the highest V_max/K_m value toward maltooctaose in the absence of chloride ion and toward maltotriose in the presence of chloride ion.     

Development and mapping of functional expressed sequence tag-derived simple sequence repeat markers in a rubber tree RRIM600 × PB217 population

Sets of polymorphic expressed sequence tag–simple sequence repeat (EST-SSR) markers from the rubber tree (Hevea brasiliensis) have been published by many researchers, but none has been specifically developed to study latex and wood yield traits. In this study, a total 10,321 rubber tree EST sequences, generated from suppression subtractive hybridization-cDNA libraries of bark and latex of high- and low-yielding clones, were used as sources for SSR searching. A total of 432 EST-SSR loci were identified and it was possible to design primer pairs for a subset of 298 EST-SSRs. The highest proportion of EST-SSRs was represented by dinucleotide repeats (46.6 %), followed by trinucleotide repeats (44.3 %). Based on BLASTX analysis, 234 ESTs (80 %) showed similarity to genes in NCBI databases and could be divided into 120 putative proteins with known function and 114 unknown proteins. To enhance the resolution of an existing linkage map from previous work on a rubber tree RRIM600 × PB217 population, 69 EST-SSR markers from the above set were tested to be integrated into the reference genetic map. The enriched map of 18 linkage groups spanned 2054.2 cM in length, showed an average genetic distance of 4.3 cM between adjacent markers, and included 63 new EST-SSR markers. The enhanced map from this study provides a basis for comparative mapping using PCR-based markers and identification of expressed genes possibly affecting important traits of interest.     

Development of 240 novel EST-SSRs in Eucalyptus L’Hérit

Simple sequence repeat (SSR) markers derived from expressed sequence tag (EST) resources provide great potential for comparative mapping, direct gene tagging of quantitative trait loci and functional diversity studies. Here we report on the development of 240 novel EST-SSRs for the important tree genus Eucalyptus L’Hérit. Of the 240 EST-SSR loci, 218 (90.8 %) were polymorphic among 12 individuals of E. grandis Hill ex Maiden, with the number of alleles per locus (N _a), observed heterozygosity (H _o), expected heterozygosity (H _e) and polymorphic information content (PIC) averaging at 5.0, 0.403, 0.598 and 0.529, respectively. High rates of cross-species/subgenus amplification were observed. The EST-SSRs developed herein would be a valuable addition of functional markers for genetics and breeding applications in a wide range of eucalypt species. The primer sequences for the 240 EST-SSRs have been deposited in the Probe database of GenBank (IDs Pr016588534–773).     

Development of microsatellite markers in red-tide causative species Prorocentrum micans (Dinophyceae)

We isolated eight polymorphic microsatellites from the red-tide causing microalgae Prorocentrum micans. These loci provide a class of highly variable genetic markers, as the number of alleles ranged from two to six with an average of 3.9, and the estimate of gene diversity was from 0.15 to 0.55 with an average of 0.41 across the eight microsatellites. We consider that these loci have a potential to reveal the genetic structure and gene flow among P. micans populations.     

基于高通量测序技术分析患病与健康虾夷扇贝(Patinopecten yessoensis)闭壳肌菌群多样性

本研究取自同一养殖海域不同浮筏养殖笼中健康和患脓包病虾夷扇贝(Patinopecten yessoensis)样品,健康虾夷扇贝样品标记为CK,患病样品分别标记为S1、S2和S3,使用Mi Seq高通量测序技术比较不同生存状态的虾夷扇贝的微生物组成与差异。结果表明:4个样品中菌群多样性表现为S3CKS1S2;4个样品中的细菌可归为24个门,主要门类有变形菌门、拟杆菌门、浮霉菌门、梭杆菌门、厚壁菌门、酸杆菌门、放线菌门、硝化螺旋菌门、绿菌门和绿弯菌门;在健康虾夷扇贝样品中,变形菌门占绝对优势,占整个菌群的95.9%,患病虾夷扇贝样品中,S1和S3中优势类群均为变形菌门,分别占整个菌群的80.4%和86.8%,S2中优势菌群为拟杆菌门,占56.9%。     

Polymerization of β-like actin from scallop adductor muscle

Scallop adductor muscle β-like isoactin differs from rabbit skeletal muscle α-actin in the rate, extent and critical concentration of polymerization. The difference is temperature- and [KCl]-dependent. In the presence of DNase I scallop actin was shown to be depolymerized more rapidly than rabbit actin. It was suggested that the polymers formed by β-actin are less stable than those formed by α-actin.     

Haploid plants from anther cultures of poplar (Populus × beijingensis)

Homozygous genotypes are valuable for genetic and genomic studies in higher plants. However, obtaining homozygous perennial woody plants using conventional breeding techniques is currently a challenge due to a long juvenile period, high heterozygosity, and substantial inbreeding depression. In vitro androgenesis has been used to develop haploid and doubled haploid plants. In the present study, we report the regeneration of haploid lines of poplar (Populus × beijingensis) via anther culture. Anthers at the uninucleate stage were induced to produce callus using three basic media. Two auxins (naphthalene acetic acid [NAA] and 2,4-dichloro-phenoxyacetic acid [2,4-D]), and two cytokinin (kinetin [KT] and 6-benzyladenine [BA]) were tested to explore the influence of plant growth regulators on callus response. H medium (Bourgin and Nitsch 1967) supplemented with 1.0 mg/L NAA and 1.0 mg/L KT induced the highest rate of callus formation. When callus obtained from anthers were subcultured in MS medium containing 1.0 mg/L BA and 0.2 mg/L NAA, followed by transfer to half-strength MS medium supplemented with indole-3-butyric acid (0.2–0.5 mg/L), the formation of regenerated plantlets increased dramatically. Inclusion of gibberellic acid (0.02–0.2 mg/L) in addition to a combination of BA (0.6 mg/L)-NAA (0.2 mg/L) in the culture medium resulted in enhanced frequency of shoot development, as well as greater internode elongation. Ploidy analysis of 580 regenerated plants, using both flow cytometry and chromosome counting, revealed 10.3 % haploid and 1.0 % triploid plantlets. The remaining plantlets were all diploid.     

The amino acid sequence of γ-crystallin (fraction II) from calf lens

The amino acid sequence of gamma-crystallin (fraction II) from calf lens has been determined; this indicates it to be a single-chain polypeptide of 165 amino acid residues.     

Cloning and nucleotide sequence of the α-galactosidase cDNA from Cyamopsis tetragonoloba (guar)

Polyadenylated mRNA was purified from the aleurone cells of Cyamopsis tetragonoloba (guar) seeds germinated for 18 h and used for the construction of a cDNA library. Clones with the -galactosidase encoding gene were identified using oligo-nucleotide mixed probes based on the NH₂ terminal amino acid sequence and on the sequence of an internal peptide. The nucleotide sequence of the cDNA clone showed that the enzyme is synthesized as a precursor with a 47 amino acid NH₂ terminal extension. This pre-sequence most likely functions to target the protein outside the aleurone cells into the endosperm. Based upon structural features, it is proposed to divide the precursor into a pre-(signal sequence) part and a glycosylated pro-part comparable with those of the yeast mat A/ factor and killer factor. A comparison of the derived amino acid sequence of this -galactosidase from plant origin revealed significant stretches of homology with respect to the amino acid sequences of the enzymes from Saccharomyces cerevisiae and from human origin but only to a minor extent compared with the -galactosidase from Escherichia coli.     

Development of SCAR Markers for Germplasm Characterisation in Olive Tree (Olea europea L.)

A set of 14 SCAR markers were developed starting from RAPD, AFLP and SAMPL analysis of several olive germplasm accessions. Eight RAPD, two AFLP and four SAMPL fragments were converted into dominant and codominant SCARs by cloning and sequencing the selected fragments. The markers obtained were evaluated on forty different olive cultivars from different Italian production areas (mainly from Liguria). The combined use of these SCARs made possible to univocally identify 26 cultivars while the remaining 14 will require the development of further markers since most of them are placed in a main group containing six genetically similar cultivars (among which Frantoio and Taggiasca) and four minor groups containing two cultivars each. A total of 31 different haplotypes were identified and the analysis of several individual plants indicated no intra-cultivar variability. Considering the SCAR polymorphism two alleles were scored for each markers with the only exception of markers IGPS3 and IGPS4 showing 4 alleles with 7 recognised groups and 5 alleles with 4 groups, respectively. Though less polymorphic in comparison with other markers like SSRs, the developed SCARs proved useful in genotype identification. In addition, they could potentially be used for breeding applications and forensic analysis. An erratum to this article is available at .     

Audiogram of the chicken (Gallus gallus domesticus) from 2 Hz to 9 kHz

The pure-tone thresholds of four domestic female chickens were determined from 2 Hz to 9 kHz using the method of conditioned suppression/avoidance. At a level of 60 dB sound pressure level (re 20 μN/m²), their hearing range extends from 9.1 Hz to 7.2 kHz, with a best sensitivity of 2.6 dB at 2 kHz. Chickens have better sensitivity than humans for frequencies below 64 Hz; indeed, their sensitivity to infrasound exceeds that of the homing pigeon. However, when threshold testing moved to the lower frequencies, the animals required additional training before their final thresholds were obtained, suggesting that they may perceive frequencies below 64 Hz differently than higher frequencies.     

Molecular basis of childhood deafness resulting from mutations in the GJB2 (connexin 26) gene

Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant bleeding disorder characterized by localized angiodysplasia. Mutations in either of two genes, endoglin or ALK-1, can cause HHT. Both genes encode putative receptors for the transforming growth factor-beta superfamily of ligands. Many mutations in each gene have been identified in HHT kindreds from around the world, and with few exceptions mutations are unique and family specific. The prevalence of HHT in the Leeward Islands of the Netherlands Antilles is possibly the highest of any geographical location. We wished to establish whether this high prevalence is due to a genetic founder effect or to multiple mutational events. HHT kindreds from the Netherlands Antilles and The Netherlands were screened for mutations in the two genes associated with HHT. Haplotype analysis of a 5-cM region on chromosome 9 flanking the endoglin gene revealed three distinct disease haplotypes in the ten Antillean families studied. Seven of these families share a splice-site mutation in exon 1 of endoglin. Two other Antillean families share a missense mutation in exon 9a of endoglin. This mutation was also found in a Dutch family that shares the same disease haplotype as the Antillean families with this mutation. Thus it appears that HHT in the Netherlands Antilles is due to a limited number of ancestral mutations in the endoglin gene, and that one of these mutations was introduced into the African slave population by a Dutch colonist. The limited scope of mutations suggests that a presymptomatic screening program for HHT would be feasible in this population.     

Nucleotide sequence of an endo-1,4-β-glucanase gene (celA) from Clostridium josui

The nucleotide sequence of a DNA fragment containing an endo-1,4-β-glucanase (EG-1) gene of Clostridium josui was determined by the dideoxy-chain termination method. The EG-1 coding sequence was an open reading frame encoding 369 amino acids, and a putative promoter sequence was located in the upstream region of the open reading frame. The N-terminal amino acid sequence of the endoglucanase (EG-1C) purified from the Escherichia coli transformant (JM103/pUCJ1) was consistent with the deduced sequence from ³⁰Val to ⁴⁴Lys. The estimated molecular weights of the precursor and the mature enzymes were 41,774 and 38,352, respectively. The region of amino acids from 61st to 335th of the enzyme revealed high homology with those of Bacillus sp. and Clostridium acetobutylicum endoglucanases.     

First record of Zombrus bicolor (Enderlein) (Hymenoptera, Braconidae, Doryctinae) in Western Europe

The finding of Zombrus bicolor (Enderlein) (Hymenoptera: Braconidae: Doryctinae) in a Tuscan vineyard of the Siena province (Italy) represents the first record of this species in western Europe. A female was captured in summer 2009 with a malaise trap located in an organic vineyard. Until this finding, the species was recorded only in the Oriental regions of continental China, Taiwan, Korea and Japan and, very recently, in the eastern and southern parts of the Palaearctic region.     

Impact on biodiversity of scallop dredging in San Matías Gulf,northern Patagonia (Argentina)

Between 1968 and 1994, Tehuelche scallop (Aequipecten tehuelchus) was fished in San Matías Gulf (Argentine Patagonia) using dredges. The catch was not sorted on board; epifaunal invertebrate bycatch and inorganic substrate removed by the dredges were landed together with scallops. Surveys were conducted in 1987 and 1997 on four fishing grounds following the same methodology to estimate the abundance of epifaunal components using catch and swept-area data. Univariate, distributional, and multivariate methods were used to analyze biodiversity and its change between surveys. There was no recorded dredging on two of the fishing grounds in the intervening period between the two surveys. Fishing effort in the order of 5,000 effective fishing hours occurred at each of the other two grounds, during that period. Multivariate analysis indicates that macrofaunal assemblages changed in each ground between 1987 and 1997, according to fishing intensity and time lapsed since last fishing action. Fishing grounds that were not dredged during the 10-year period showed no significant changes in species composition and dominance. The results support the hypothesis that dredging causes high levels of disturbance that affect the whole benthic ecosystem. Community “recovery” seems to be a non-stationary process that may not reverse the system to pre-harvest conditions, but lead instead to a sequence of new states with faunistic changes governed by colonization and slow rebuilding of habitat complexity. Handling editor: K. Martens     

Diversity of actinomycetes isolated from Challenger Deep sediment (10,898 m) from the Mariana Trench

Thirty-eight actinomycetes were isolated from sediment collected from the Mariana Trench (10,898 m) using marine agar and media selective for actinomycetes, notably raffinose-histidine agar. The isolates were assigned to the class Actinobacteria using primers specific for members of this taxon. The phylogenetic analysis based on 16S rRNA gene sequencing showed that the isolates belonged to the genera Dermacoccus, Kocuria, Micromonospora, Streptomyces, Tsukamurella and Williamsia. All of the isolates were screened for genes encoding nonribosomal peptide and polyketide synthetases. Nonribosomal peptide synthetase sequences were detected in more than half of the isolates and polyketide synthases type I (PKS-I) were identified in five out of 38 strains. The Streptomyces isolates produced several unusual secondary metabolites, including a PKS-I associated product. In initial testing for piezotolerance, the Dermacoccus strain MT1.1 grew at elevated hydrostatic pressures.     

Development and characterization of microsatellite loci from the Great White Pelican (Pelecanus onocrotalus) and widespread application to other members of the Pelecanidae

We describe the isolation of 10 microsatellite loci from the Great White Pelican using an enrichment protocol. All loci were variable with the number of alleles ranging from 2 to 19, and observed heterozygosity ranging from 0.261 to 0.913. Two loci were out of Hardy–Weinberg equilibrium, although in each case this was restricted to one of the two populations screened. None of the loci were linked. Eight to 10 loci PCR-amplified in three related species: Brown, American White and Pink-backed Pelicans. These loci should prove to be widely applicable to studies of phylogeography and demography in pelicans globally.