Development of a rapid, sensitive and specific diagnostic assay for fish Aquareovirus based on RT-PCR

A rapid, sensitive and highly specific detection method for Aquareovirus based on reverse-transcription polymerase chain reaction (RT-PCR) was developed. Based on multiple sequence alignment of the cloned sequences of a local isolates, the Threadfin reovirus (TFV) and Guppy reovirus (GPV) with Grass carp reovirus (GCRV), a pair of degenerate primers was selected carefully and synthesized. Using this primer combination, only one specific product, approximately 450 bp in length was obtained when RT-PCR was carried out using the genomic double-stranded RNA (dsRNA) of TFV, GPV and GCRV. Similar results were also obtained when Chum salmon reovirus (CSRV) and Striped bass reovirus (SBRV) dsRNA were used as templates. No products were observed when nucleic acids other than the dsRNA of the aquareoviruses described above were used as RT-PCR templates. This technique could detect not only TFV but also GPV and GCRV in low titer virus-infected cell cultured cells. Furthermore, this method has also been shown to be able to diagnose GPV-infected guppy (Poecilia reticulata) that exhibit clinical symptoms as well as GPV-carrier guppy. Collectively, these results showed that the RT-PCR amplification method using specific degenerate primers described below is very useful for rapid and accurate detection of a variety of aquareovirus strains isolated from different host species and origin. (C) 2004 Elsevier B.V. All rights reserved.

Intraspecific phylogeography of Carchesium polypinum (Peritrichia, Ciliophora) from China, inferred from 18S-ITS1-5.8S ribosomal DNA

Based on the variation of site 34, 46, 241, 305 and 322 in the 18S-ITS1 rDNA sequence, 19 Carchesium polypinum populations collected from eight provinces of China were separated into northern and southern population along the delineation between the Yangtze River and the Pearl River. This geographic distribution pattern of Carchesium polypinum maybe results from two factors: the vicariance resulting from the formation of the delineation between the Pearl River and the Yangtze River accompanied with the uplift of Qinghai-Xizang Plateau, and the different dispersal paths of C. polypinum affected by the climate.

Cross-species amplification in silver carp and bighead carp with microsatellite primers of common carp

About a third of microsatellite primers designed for common carp (Cyprinus carpio) was successfully amplified in silver carp (Hypophthalmichthys molitrix) and bighead carp (Aristichthys nobilis). These markers, inherited in Mendelian mode, are of potential applications in cypinid genetics.

The molecular characterization of RNA segment S9 of grass carp hemorrhage virus (GCHV), an aquareovirus

Although reovirus infection is one of the major virus diseases of grass carp in China, the available knowledge on the structure and function of genes and proteins of the virus is limited. The complete sequence of the S9 genome segment of grass carp hemorrhage virus (GCHV) was determined. The segment consists of 1130 nucleotides and has a large open reading frame (ORF) encoding a protein of 352 amino acids with predicted molecular mass of 37.7 kDa. Amino acid sequence comparison revealed that the deduced protein encoded by GCHV S9 is closely related to the sigma NS proteins of mammalian reovirus (MRV) and avian reovirus (ARV). Secondary structure analysis displayed that the form of alpha -helices (40.1%) and beta -sheets (49.4%) are the richest two contents in the protein encoded by S9, and this protein is predicted to be a nonstructural protein. (C) 2001 Elsevier Science B.V. All rights reserved.

A detection method for grass carp hemorrhagic virus (GCHV) based on a reverse transcription polymerase chain reaction

A rapid, sensitive and highly specific detection method for grass carp hemorrhagic virus (GCHV) based on a reverse transcription-polymerase chain reaction (RT-PCR) has been developed. Two pairs of PCR primers were synthesized according to the cloned cDNA sequences of the GCHV-861 strain. For each primer combination, only one specific major product was obtained when amplification was performed by using the genomic dsRNA of GCHV-861 strain. The lengths of their expected products were 320 and 223 bp, respectively. No products were obtained when nucleic acids other than GCHV-861 genomic RNA were used as RT-PCR templates. To assess the sensitivity of the method, dilutions of purified GCHV-861 dsRNA total genome (0.01 pg up to 1000 pg) were amplified and quantities of as little as 0.1 pg of purified dsRNA were detectable when the amplification product was analyzed by 1.5% agarose gel electrophoresis. This technique could detect GCHV-861 not only in infected cell culture fluids, but also in infected grass carp Ctenopharyngodon idellus and rare minnow Gobiocypris rarus with or without hemorrhagic symptoms. The results show that the RT-PCR amplification method is useful for the direct detection of GCHV.

PURIFICATION AND CHARACTERIZATION OF A KINASE SPECIFIC FOR THE SERINE-RICH AND ARGININE-RICH PRE-MESSENGER-RNA SPLICING FACTORS

Members of the SR family of pre-mRNA splicing factors are phosphoproteins that share a phosphoepitope specifically recognized by monoclonal antibody (mAb) 104. Recent studies have indicated that phosphorylation may regulate the activity and the intracellular localization of these splicing factors. Here, we report the purification and kinetic properties of SR protein kinase 1 (SRPK1), a kinase specific for SR family members. We demonstrate that the kinase specifically recognizes the SR domain, which contains serine/arginine repeats. Previous studies have shown that dephosphorylated SR proteins did not react with mAb 104 and migrated faster in SDS gels than SR proteins from mammalian cells. We show that SRPK1 restores both mobility and mAB 104 reactivity to a SR protein SF2/ASF (splicing factor 2/alternative splicing factor) produced in bacteria, suggesting that SRPK1 is responsible for the generation of the mAb 104-specific phosphoepitope in vivo. Finally, we have correlated the effects of mutagenesis in the SR domain of SF2/ASF on splicing with those on phosphorylation of the protein by SRPK1, suggesting that phosphorylation of SR proteins is required for splicing.

Room-temperature highly anisotropic ferromagnetism and uniaxial spin amplification in (In,Mn)As quantum dots

The hole-mediated ferromagnetism in (In,Mn)As quantum dots is investigated using the k center dot p method and the mean field model. It is found that the (In,Mn)As quantum dot can be ferromagnetic at room temperature when there is one hole in the dot. For the spherical quantum dots, the Curie temperature decreases as the diameter increases, and increases as the effective composition of magnetic ions increases. It is interesting to find that the (In,Mn)As oblate quantum dot has highly anisotropic Zeeman splitting and ferromagnetism due to the spin-orbit coupling effect, which can be used as an uniaxial spin amplifier. (c) 2008 American Institute of Physics.

Ultra-broadband optical parametric chirped-pulse amplification by matching of both group-velocity and pulse-front

Ultra-broadband optical parametric chirped-pulse amplification is analyzed based the compensation of phase-mismatch, which is achieved by matching of both group-velocity and pulse-front between signal and idler by the combination of the noncollinear-phase-match and pulse-front-tilt. The results show exactly matching of both group-velocity and pulse-front is the important criterion for constructing an UBOPCPA. Its general model is developed, in which the group velocities, noncollinear angles. spatial walk-off angles, linear angular spectral dispersion coefficients and pulse-front tilted angles are suitably linked to each other. Finally, specific numerical calculations and simulations are presented for beta-barium borate OPCPA with type-1 noncollinear angularly dispersed geometry. (C) 2005 Elsevier B.V. All rights reserved.

Experimental and theoretical analysis of nondegenerate ultrabroadband chirped pulse optical parametric amplification

Experimental investigations of nondegenerate ultrabroadband chirped pulse optical parametric amplification have been carried out. The general mathematical expressions for evaluating parametric bandwidth, gain and gain bandwidth for arbitrary three-wave mixing parametric amplifiers are presented. In our experiments, a type-I noncollinear phase-matched optical parametric amplifier based on lithium triborate, which was pumped by a 5-ns second harmonic pulses from a Q-switched Nd:YAG operating at 10 Hz, seeded by a 14-fs Ti:sapphire laser at 800 nm, was presented. The 0.85 nJ energy of input chirped signal pulse with 57-FWHM has been amplified to 3.1 muJ at pump intensity 3 GW/cm(2), the corresponding parametric gain reached 3.6 x 10(3), the 53 nm-FWHM gain spectrum bandwidth of output signal has been obtained. The large gain and broad gain bandwidth, which have been confirmed experimentally, provide great potentials to amplify efficiently the broad bandwidth femtosecond light pulses to generate new extremes in power, intensity, and pulse duration using optical parametric chirped pulse amplifiers pumped by powerful nanosecond systems.

Investigation of spectral bandwidth of optical parametric amplification

The spectral bandwidth of three-wave-mixing optical parametric amplification has been investigated. A general mathematical model for evaluating the spectral bandwidth of optical parametric amplification is developed with parametric bandwidth and gain bandwidth via three-wave noncollinear interactions. The spectral bandwidth is determined by expanding the wave-vector mismatch in a Taylor series and retaining terms through second order. The model takes into account the effects of crystal length, noncollinear angle, group velocity, group-velocity dispersion and gain coefficient. The relation between parametric bandwidth and gain bandwidth is clearly defined. The model is applied to a BBO OPA, a LBO OPA and a CLBO OPA.

Compact and efficient triple-pass optical parametric chirped pulse amplification

Compact and efficient triple-pass optical parametric chirped pulse amplification in a single crystal has been demonstrated. The signal was triple-pass amplified in a single nonlinear crystal by a nanosecond pump pulse. The first-pass optical parametric amplification is completely phase matched in the plane of the maximum effective nonlinearity, and the other two passes work symmetrically near to the first-pass optical parametric amplification plane. This architecture efficiently increases the overall gain, overcomes the optical parametric fluorescence, and clearly simplifies the amplification scheme.

Stacking chirped pulse optical parametric amplification

Stacking chirped pulse optical parametric amplification based on a home-built Yb(3+)-doped mode-locked fiber laser and an all-fiber pulse stacker has been demonstrated. Energic 11 mJ shaped pulses with pulse duration of 2.3 ns and a net total gain of higher than 1.1 x 10(7) at fluctuation less than 2% rms are achieved by optical parametric amplification pumped by a Q-switched Nd:YAG frequency-doubled laser, which provides a simple and efficient amplification scheme for temporally shaped pulses by stacking chirped pulse. (C) 2009 Elsevier B.V. All rights reserved.