Development and validation of molecular markers for characterization of Boehmeria nivea var. nivea and Boehmeria nivea var. tenacissima

<p>Abstract</p> <p>Background</p> <p>The root of <it>Boehmeria </it>spp (ramie) is a hepatoprotective Chinese herbal medicine. Medicinal properties vary between <it>Boehmeria nivea </it>var. <it>nivea </it>and <it>Boehmeria nivea </it>var. <it>tenacissima</it>, which are local species found in Taiwan. As commercial preparations may use either species, there is a need for a rapid and simple assay to identify variants for quality control.</p> <p>Methods</p> <p>Four methods were developed and tested for their applicability in differentiating the two species. These methods were random amplified polymorphic DNA (RAPD); sequence characterized amplified regions (SCAR); single nucleotide polymorphisms (SNP) and cleaved amplified polymorphic sequences (CAPS).</p> <p>Results</p> <p>Three RAPD markers were developed that produced unique bands in <it>B. nivea </it>var. <it>tenacissima </it>and <it>B. nivea </it>var. <it>nivea</it>. Based on sequenced RAPD bands, one SCAR marker was developed that produced a single DNA band in <it>B. nivea </it>var. <it>nivea</it>. Two SNP markers differentiated between <it>B. nivea </it>var. <it>nivea </it>and <it>B. nivea </it>var. <it>tenacissima </it>based on single nucleotide substitutions. A pair of CAPS oligonucleotides was developed by amplifying a 0.55-kb DNA fragment that exhibited species-specific digestion patterns with restriction enzymes <it>Alf </it>III and <it>Nde </it>I. Consistent results were obtained with all the four markers on all tested <it>Boehmeria </it>lines.</p> <p>Conclusion</p> <p>The present study demonstrates the use of the RAPD, SCAR, SNP and CAPS markers for rapid identification of two closely related <it>Boehmeria </it>species.</p

Development of a fluorescent quantitative real-time polymerase chain reaction assay for the detection of Goose parvovirus in vivo

<p>Abstract</p> <p>Background</p> <p>Goose parvovirus (GPV) is a <it>Dependovirus </it>associated with latent infection and mortality in geese. Currently, it severely affects geese production worldwide. The objective of this study was to develop a fluorescent quantitative real-time polymerase chain reaction (PCR) (FQ-PCR) assay for fast and accurate quantification of GPV DNA in infected goslings, which can aid in the understanding of the regular distribution pattern and the nosogenesis of GPV in vivo.</p> <p>Results</p> <p>The detection limit of the assay was 2.8 × 10¹standard DNA copies, with a sensitivity of 3 logs higher than that of the conventional gel-based PCR assay targeting the same gene. The real-time PCR was reproducible, as shown by satisfactory low intraassay and interassay coefficients of variation.</p> <p>Conclusion</p> <p>The high sensitivity, specificity, simplicity, and reproducibility of the GPV fluorogenic PCR assay, combined with a high throughput, make this method suitable for a broad spectrum of GPV etiology-related applications.</p

Sodium azide mutagenesis resulted in a peanut plant with elevated oleate content

Screening of peanut seeds resulting from 0.39% sodium azide treatment with NIRS calibration equation for bulk seed samples identified a plant with more than 60% oleate. Oleate content in individual seeds of the plant, as predicted by NIRS calibration equation for intact single peanut seeds, ranged from 50.05% ~ 68.69%. Three seeds with &gt;60% oleate thus identified were further confirmed by gas chromatography. Multiple sequence alignments of the FAD2B gene from Huayu 22 (wild type) and peanut seeds with elevated oleate (mutant type) revealed a C281T transition in the coding region causing an I94T substitution in the oleoyl-PC desaturase, which may be responsible for reduction in the enzyme activity

Molecular and clinical analyses of 84 patients with tuberous sclerosis complex

BACKGROUND: Tuberous sclerosis complex (TSC) is an autosomal dominant disease characterized by the development of multiple hamartomas in many internal organs. Mutations in either one of 2 genes, TSC1 and TSC2, have been attributed to the development of TSC. More than two-thirds of TSC patients are sporadic cases, and a wide variety of mutations in the coding region of the TSC1 and TSC2 genes have been reported. METHODS: Mutational analysis of TSC1 and TSC2 genes was performed in 84 Taiwanese TSC families using denaturing high-performance liquid chromatography (DHPLC) and direct sequencing. RESULTS: Mutations were identified in a total of 64 (76 %) cases, including 9 TSC1 mutations (7 sporadic and 2 familial cases) and 55 TSC2 mutations (47 sporadic and 8 familial cases). Thirty-one of the 64 mutations found have not been described previously. The phenotype association is consistent with findings from other large studies, showing that disease resulting from mutations to TSC1 is less severe than disease due to TSC2 mutation. CONCLUSION: This study provides a representative picture of the distribution of mutations of the TSC1 and TSC2 genes in clinically ascertained TSC cases in the Taiwanese population. Although nearly half of the mutations identified were novel, the kinds and distribution of mutation were not different in this population compared to that seen in larger European and American studies

Mechanism for controlling the monomer–dimer conversion of SARS coronavirus main protease

[[abstract]]The Severe acute respiratory syndrome coronavirus (SARS-CoV) main protease (Mpro) cleaves two virion polyproteins (pp1a and pp1ab); this essential process represents an attractive target for the development of anti-SARS drugs. The functional unit of Mpro is a homodimer and each subunit contains a His41/Cys145 catalytic dyad. Large amounts of biochemical and structural information are available on Mpro; nevertheless, the mechanism by which monomeric Mpro is converted into a dimer during maturation still remains poorly understood. Previous studies have suggested that a C-terminal residue, Arg298, interacts with Ser123 of the other monomer in the dimer, and mutation of Arg298 results in a monomeric structure with a collapsed substrate-binding pocket. Interestingly, the R298A mutant of Mpro shows a reversible substrate-induced dimerization that is essential for catalysis. Here, the conformational change that occurs during substrate-induced dimerization is delineated by X-ray crystallography. A dimer with a mutual orientation of the monomers that differs from that of the wild-type protease is present in the asymmetric unit. The presence of a complete substrate-binding pocket and oxyanion hole in both protomers suggests that they are both catalytically active, while the two domain IIIs show minor reorganization. This structural information offers valuable insights into the molecular mechanism associated with substrate-induced dimerization and has important implications with respect to the maturation of the enzyme.[[notice]]補正完畢[[incitationindex]]SCI[[booktype]]電子

Characterization of Aniline Tetramer by MALDI TOF Mass Spectrometry upon Oxidative and Reductive Cycling.

By combining electrochemical experiments with mass spectrometric analysis, it is found that using short chain oligomers to improve the cycling stability of conducting polymers in supercapacitors is still problematic. Cycling tests via cyclic voltammetry over a potential window of 0 to 1.0 V or 0 to 1.2 V in a two-electrode device configuration resulted in solid-state electropolymerization and chain scission. Electropolymerization of the aniline tetramer to generate long chain oligomers is shown to be possible despite the suggested decrease in reactivity and increase in intermediate stability with longer oligomers. Because aniline oligomers are more stable towards reductive cycling when compared to oxidative cycling, future conducting polymer/oligomer-based pseudocapacitors should consider using an asymmetric electrode configuration

Genome-Wide Association Study Identifies ALDH7A1 as a Novel Susceptibility Gene for Osteoporosis

Osteoporosis is a major public health problem. It is mainly characterized by low bone mineral density (BMD) and/or low-trauma osteoporotic fractures (OF), both of which have strong genetic determination. The specific genes influencing these phenotypic traits, however, are largely unknown. Using the Affymetrix 500K array set, we performed a case-control genome-wide association study (GWAS) in 700 elderly Chinese Han subjects (350 with hip OF and 350 healthy matched controls). A follow-up replication study was conducted to validate our major GWAS findings in an independent Chinese sample containing 390 cases with hip OF and 516 controls. We found that a SNP, rs13182402 within the ALDH7A1 gene on chromosome 5q31, was strongly associated with OF with evidence combined GWAS and replication studies (P = 2.08×10−9, odds ratio = 2.25). In order to explore the target risk factors and potential mechanism underlying hip OF risk, we further examined this candidate SNP's relevance to hip BMD both in Chinese and Caucasian populations involving 9,962 additional subjects. This SNP was confirmed as consistently associated with hip BMD even across ethnic boundaries, in both Chinese and Caucasians (combined P = 6.39×10−6), further attesting to its potential effect on osteoporosis. ALDH7A1 degrades and detoxifies acetaldehyde, which inhibits osteoblast proliferation and results in decreased bone formation. Our findings may provide new insights into the pathogenesis of osteoporosis

Foxp2 controls synaptic wiring of corticostriatal circuits and vocal communication by opposing Mef2c

Cortico-basal ganglia circuits are critical for speech and language and are implicated in autism spectrum disorder, in which language function can be severely affected. We demonstrate that in the mouse striatum, the gene Foxp2 negatively interacts with the synapse suppressor gene Mef2c. We present causal evidence that Mef2c inhibition by Foxp2 in neonatal mouse striatum controls synaptogenesis of corticostriatal inputs and vocalization in neonates. Mef2c suppresses corticostriatal synapse formation and striatal spinogenesis, but can itself be repressed by Foxp2 through direct DNA binding. Foxp2 deletion de-represses Mef2c, and both intrastriatal and global decrease of Mef2c rescue vocalization and striatal spinogenesis defects of Foxp2-deletion mutants. These findings suggest that Foxp2-Mef2C signaling is critical to corticostriatal circuit formation. If found in humans, such signaling defects could contribute to a range of neurologic and neuropsychiatric disorders.National Institutes of Health (U.S.) (Grant R37 HD028341)Eunice Kennedy Shriver National Institute of Child Health and Human Development (U.S.) (Award R37 HD028341