Homogeneous genetic structure and variation in tree architecture of <Emphasis Type="Italic">Larix kaempferi</Emphasis> along altitudinal gradients on Mt. Fuji

Variations in tree architecture and in the genetic structure of Larix kaempferi on Mt. Fuji were surveyed along altitudinal gradients using 11 nSSR loci. In total, 249 individuals from six populations along three trails at altitudes ranging from approximately 1,300 to 2,700 m were investigated. Gradual changes in tree architecture with increasing elevation, from erect trees to flag trees and krummholz mats, were observed in the high-altitude populations (>2,000 m) on all trails. These findings suggest that tree architecture is correlated with the severe environmental conditions associated with increasing elevation, such as strong winds. In contrast to obvious variations in tree architecture, the genetic diversity of populations along the trails was almost uniform (H _E = 0.717–0.762) across the altitudinal range. The results of the AMOVA and STRUCTURE analyses, and the analysis for isolation by distance pattern, suggest homogeneous genetic structuring across all populations on Mt. Fuji, while the pairwise F _ST showed barriers to gene flow between altitudinal populations that were demarcated as high- or low-altitude populations by Abies-Tsuga forest. Although the evergreen coniferous forests on the mountainside may hinder gene flow, this may be explained by the long-distance seed dispersal of the Japanese larch and/or a short population history resulting from eruptions or slush avalanches, although evergreen coniferous forests on the mountainside may hinder gene flow.     

Salt-dependent thermo-reversible α-amylase: cloning and characterization of halophilic α-amylase from moderately halophilic bacterium, <Emphasis Type="Italic">Kocuria varians</Emphasis>

A moderately halophilic bacterium, Kocuria varians, was found to produce active α-amylase (K. varians α-amylase (KVA)). We have observed at least six different forms of α-amylase secreted by this bacterium into the culture medium. Characterization of these KVA forms and cloning of the corresponding gene revealed that KVA comprises pre-pro-precursor form of α-amylase catalytic domain followed by the tandem repeats, which show high similarity to each other and to the starch binding domain (SBD) of other α-amylases. The observed six forms were most likely derived by various processing of the protein product. Recombinant KVA protein was successfully expressed in Escherichia coli as a fusion protein and was purified with affinity chromatography after cleavage from fusion partner. The highly acidic amino acid composition of KVA and the highly negative electrostatic potential surface map of the modeled structure strongly suggested its halophilic nature. Indeed, KVA showed distinct salt- and time-dependent thermal reversibility: when α-amylase was heat denatured at 85°C for 3 min in the presence of 2 M NaCl, the activity was recovered upon incubation on ice (50% recovery after 15 min incubation). Conversely, KVA denatured in 0.1 M NaCl was not refolded at all, even after prolonged incubation. KVA activity was inhibited by proteinaceous α-amylase inhibitor from Streptomyces nitrosporeus, which had been implicated to inhibit only animal α-amylases. KVA with putative SBD regions was found to digest raw starch.     

Sequence analysis of the α-galactosidase <Emphasis Type="Italic">MEL</Emphasis> gene governing the efficient production of ethanol from raffinose-rich molasses in the yeast <Emphasis Type="Italic">Lachancea thermotolerans</Emphasis>

The yeast Lachancea thermotolerans, formerly Kluyveromyces thermotolerans, was tested for the ethanol fermentation of raffinose-rich molasses. Two melibiose-fermenting strains, NBRC 10066 and NBRC 10067, produced more ethanol than eight other strains. The concentration of ethanol synthesized by NBRC 10066 was slightly higher than that by NBRC 10067, probably on the basis of the expression of α-galactosidase. The regions corresponding to the α-galactosidase MEL1 gene of Saccharomyces cerevisiae were amplified. The nucleotide sequences of the two genes designated as MELth1 and MELth2 revealed single open reading frames of 1,416 bp encoding 472 amino acids but differed from each other in one base that converted the amino acid composition. The sequences of the 5′-upstream region from −1 to −515 of the two genes are identical except for one base.     

Adjusting estimative prediction limits

This note presents a direct adjustment of the estimative predictionlimit to reduce the coverage error from a target value to third-orderaccuracy. The adjustment is asymptotically equivalent to thoseof Barndorff-Nielsen & Cox (1994, 1996) and Vidoni (1998).It has a simpler form with a plug-in estimator of the coverageprobability of the estimative limit at the target value.     

Synthesis of highly ordered cellulose II in vitro using cellodextrin phosphorylase

Synthesis of cellulose in vitro is expected to afford tailor-made cellulosic materials with highly homogeneous structure compared to natural cellulosic materials. Here we report the enzymatic synthesis of cellulose II with high crystallinity from glucose and α-glucose 1-phosphate (αG1P) by cellodextrin phosphorylase (CDP). Although glucose had been believed not to act as a glucosyl acceptor of CDP, a significant amount of insoluble cellulose was precipitated without accumulation of soluble cello oligosaccharides when glucose was mixed with αG1P and CDP. This phenomenon can be explained in terms of the large difference in acceptor reactivity between glucose and cello oligosaccharides. ¹H NMR spectrometric analysis revealed that this insoluble cellulose had an average degree of polymerization (DP) of nine. TEM observation, together with electron and X-ray diffraction studies, indicated that the insoluble cellulose formed platelet-shaped single lamellar crystals of cellulose II, several μm in length and several hundred nm in width; this is large compared to reported cellulose crystals. The thickness of the lamellar crystal is 4.5 nm, which is equivalent to a chain length of a cello oligosaccharide with DP nine and is consistent with the ¹H NMR spectroscopic results. These results suggest that cello oligosaccharides having an average DP of nine are synthesized in vitro by CDP when glucose is used as an acceptor, and the product forms highly crystalline cellulose II when it precipitates.     

Evolution of Eastern Pacific Warm Pool and upwelling processes since the middle Miocene based on analysis of radiolarian assemblages: Response to Indonesian and Central American Seaways

A quantitative radiolarian study at Ocean Drilling Program Site 1241 in the eastern tropical Pacific enables us to reconstruct paleoceanographic changes that occurred since the latest middle Miocene. Today, this site is located just under the Eastern Pacific Warm Pool (EPWP). Based on the abundance variations of radiolarian characteristic species which are indicators of upwelling and thermocline changes, it is suggested that three notable changes occurred at 10.6, 9.8, and 4.2 Ma in the region. Four distinct periods of oceanographic conditions bounded by these notable changes were characterized on the basis of the following: (1) stratified seawater (12.0 to 10.6 Ma); (2) a shallowing of the thermocline and an increasing of upwelling (10.6 to 9.8 Ma); (3) significant inflow of warm water to the eastern tropical Pacific caused by an intensified Northern Equatorial Countercurrent (NECC), resulting in the formation of EPWP (9.8 to 4.2 Ma); and (4) the reduction of the EPWP and the NECC, and an increase in upwelling (4.2 to 0 Ma). The timing of these paleoceanographic events indicated the strong relations with the opening and closing of the Indonesian and Central American (Panama) Seaways. The reduction of the EPWP (this study) and the deepening of the thermocline in western Pacific at about 4.2 Ma (Cannariato and Ravelo, 1997; Chaisson and Ravelo, 2000) indicated a change from a state resembling El Niño in the late Miocene and the early Pliocene time to a state resembling La Niña by the late Pliocene.