Small RNAs in tomato fruit and leaf development

Tomato fruit and leaf development offers excellent systems to study the evolution of gene regulation underlying development of different organs. We have identified over 350 and 700 small RNAs from tomato fruit and leaf, respectively. Except for conserved microRNAs, more than 90% of the small RNAs are unique to tomato. We confirmed expression of some conserved as well as novel putative microRNAs by Northern hybridization. Our results help form a basis for comparative studies on how small RNA-mediated gene expression has contributed to the evolution of common and distinct developmental pathways of fruits and leaves. We have established a website (http://ted.bti.cornell.edu/digital/sRNA/) for public access to all of our small RNA sequences, their expression patterns in respective tissues, and their matching genes or predicted target genes in a searchable manner.     

Virtual electrodes and the induction of fibrillation in Langendorff-perfused rabbit ventricles: the role of intracellular calcium

A strong premature electrical stimulus (S(2)) induces both virtual anodes and virtual cathodes. The effects of virtual electrodes on intracellular Ca(2+) concentration ([Ca(2+)](i)) transients and ventricular fibrillation thresholds (VFTs) are unclear. We studied 16 isolated, Langendorff-perfused rabbit hearts with simultaneous voltage and [Ca(2+)](i) optical mapping and for vulnerable window determination. After baseline pacing (S(1)), a monophasic (10 ms anodal or cathodal) or biphasic (5 ms-5 ms) S(2) was applied to the left ventricular epicardium. Virtual electrode polarizations and [Ca(2+)](i) varied depending on the S(2) polarity. Relative to the level of [Ca(2+)](i) during the S(1) beat, the [Ca(2+)](i) level 40 ms after the onset of monophasic S(2) increased by 36+/-8% at virtual anodes and 20+/-5% at virtual cathodes (P<0.01), compared with 25+/-5% at both virtual cathode-anode and anode-cathode sites for biphasic S(2). The VFT was significantly higher and the vulnerable window significantly narrower for biphasic S(2) than for either anodal or cathodal S(2) (n=7, P<0.01). Treatment with thapsigargin and ryanodine (n=6) significantly prolonged the action potential duration compared with control (255+/-22 vs. 189+/-6 ms, P<0.05) and eliminated the difference in VFT between monophasic and biphasic S(2), although VFT was lower for both cases. We conclude that virtual anodes caused a greater increase in [Ca(2+)](i) than virtual cathodes. Monophasic S(2) is associated with lower VFT than biphasic S(2), but this difference was eliminated by the inhibition of the sarcoplasmic reticulum function and the prolongation of the action potential duration. However, the inhibition of the sarcoplasmic reticulum function also reduced VFT, indicating that the [Ca(2+)](i) dynamics modulate, but are not essential, to ventricular vulnerability.     

Expressed sequence tag analysis of Antarctic hairgrass Deschampsia antarctica from King George Island, Antarctica

Deschampsia antarctica is the only monocot that thrives in the tough conditions of the Antarctic region. It is an invaluable resource for the identification of genes associated with tolerance to various environmental pressures. In order to identify genes that are differentially regulated between greenhouse-grown and Antarctic field-grown plants, we initiated a detailed gene expression analysis. Antarctic plants were collected and greenhouse plants served as controls. Two different cDNA libraries were constructed with these plants. A total of 2,112 cDNA clones was sequenced and grouped into 1,199 unigene clusters consisting of 243 consensus and 956 singleton sequences. Using similarity searches against several public databases, we constructed a functional classification of the ESTs into categories such as genes related to responses to stimuli, as well as photosynthesis and metabolism. Real-time PCR analysis of various stress responsive genes revealed different patterns of regulation in the different environments, suggesting that these genes are involved in responses to specific environmental factors.     

Red to red - the marine bacterium Hahella chejuensis and its product prodigiosin for mitigation of harmful algal blooms

Harmful algal blooms (HABs), commonly called red tides, are caused by some toxic phytoplanktons, and have made massive economic losses as well as marine environmental disturbances. As an effective and environment-friendly strategy to control HAB outbreaks, biological methods using marine bacteria capable of killing the harmful algae or algicidal extracellular compounds from them have been given attention. A new member of the gamma-Proteobacteria, Hahella chejuensis KCTC 2396, was originally isolated from the Korean seashore for its ability to secrete industrially useful polysaccharides, and was characterized to produce a red pigment. This pigment later was identified as an alkaloid compound, prodigiosin. During the past several decades, prodigiosin has been extensively studied for its medical potential as immunosuppressants and antitumor agents, owing to its antibiotic and cytotoxic activities. The lytic activity of this marvelous molecule against Cochlodinium polykrikoides cells at very low concentrations (1 ppb) was serendipitously detected, making H. chejuensis a strong candidate among the biological agents for HAB control. This review provides a brief overview of algicidal marine bacteria and their products, and describes in detail the algicidal characteristics, biosynthetic process, and genetic regulation of prodigiosin as a model among the compounds active against red-tide organisms from the biochemical and genetic viewpoints.     

Removal of heavy metals by an enriched consortium

An enriched consortium obtained from lake-sediment was developed for the removal of heavy metals such as Cu, Pb, Cr, Ni, and Zn from heavy metal-contaminated water. The removal efficiency of heavy metals in a shaking condition was generally higher than that in the static state. After the fifteenth enrichment with assorted heavy metals, the removal efficiencies in the shaking and static condition at an average concentration of 100 mg/L of each heavy metal were approximately 99 approximately 100% and 95 approximately 100%, respectively, depending on the type of heavy metal. An aerobically grown, pure culture isolated from an enriched culture was analyzed by 16S rRNA sequencing and identified as Ralstonia sp. HM-1. This strain was found to remove various heavy metals with an efficiency of approximately 97 approximately 100% at an average concentration of 200 mg/L of each heavy metal.     

Rapid sexing of preimplantation bovine embryo using consecutive and multiplex polymerase chain reaction (PCR) with biopsied single blastomere.

The objective of this study was to establish a rapid and reliable PCR method for the sexing of 8- to 16-cell stage bovine embryos. The BOV97M and bovine 1.715 satellite DNA sequences were selected for amplification of male- and bovine-specific DNA, respectively. But the unequal number of copies of these two repetitive sequences required some modification of the multiplex PCR method. In consecutive and multiplex PCR, the first 10 PCR cycles were done with male-specific primer followed by an additional 23 cycles with bovine-specific primer. In this PCR method, the appearance of male- and bovine-specific bands was independent of the DNA concentration. This PCR method was applied successfully using groups of 8, 4, 2, and 1 blastomeres dissociated from the embryos, and the sexing efficiency was 100.0, 96.3, 94.3 and 92.1%, respectively. The coincident rate of sex determination between biopsied single blastomere and matched blastocyst was 90.0%. Therefore the developmental potential from 8- to 16-cell stage embryos to the blastocyst stage was not significantly different (P>0.2) for intact embryo (42.3%) than for demi-embryos (53.8%), suggesting that trauma to the demi-embryo caused by single-blastomere aspiration using a bevelled micropipette was very small. In conclusion, we developed a rapid (within 2 hours) and effective PCR method for the sexing of 8- to 16-cell stage bovine embryos using a single blastomere.     

Fine-scale distribution of ectomycorrhizal fungi colonizing Tsuga diversifolia seedlings growing on rocks in a subalpine Abies veitchii forest

Numerous species of ectomycorrhizal (ECM) fungi coexist under the forest floor. To explore the mechanisms of coexistence, we investigated the fine-scale distribution of ECM fungal species colonizing root tips in the root system of Tsuga diversifolia seedlings in a subalpine forest. ECM root tips of three seedlings growing on the flat top surface of rocks were sampled after recording their positions in the root system. After the root tips were grouped by terminal-restriction fragment length polymorphism (T-RFLP) analysis of ITS rDNA, the fungal species representing each T-RFLP group were identified using DNA sequencing. Based on the fungal species identification, the distribution of root tips colonized by each ECM fungus was mapped. Significant clustering of root tips was estimated for each fungal species by comparing actual and randomly simulated distributions. In total, the three seedlings were colonized by 40 ECM fungal species. The composition of colonizing fungal species was quite different among the seedlings. Twelve of the 15 major ECM fungal species clustered significantly within a few centimeters. Some clusters overlapped or intermingled, while others were unique. Areas with high fungal species diversity were also identified in the root system. In this report, the mechanisms underlying generation of these ECM root tip clusters in the root system are discussed.