Site-directed gene integration in transgenic zebrafish mediated by cre recombinase using a combination of mutant Lox sites

With current gene-transfer techniques in fish, insertion of DNA into the genome occurs randomly and in many instances at multiple sites. Associated position effects, copy number differences, and multiple gene interactions make gene expression experiments difficult to interpret and fish phenotype less predictable. To meet different fish engineering needs, we describe here a gene targeting model in zebrafish. At first, four target zebrafish lines, each harboring a single genomic lox71 target site, were generated by zebrafish transgenesis. The zygotes of transgenic zebrafish lines were coinjected with capped Cre mRNA and a knockin vector pZklox66RFP. Site-specific integration event happened from one target zebrafish line. In this line two integrant zebrafish were obtained from more than 80,000 targeted embryos (integrating efficiency about 10(-4) to 10(-5)) and confirmed to have a sole copy of the integrating DNA at the target genome site. Genomic polymerase chain reaction analysis and DNA sequencing verified the correct gene target events where lox71 and lox66 have accurately recombined into double mutant lox72 and wild-type loxP. Each integrant zebrafish chosen for analysis harbored the transgene rfp at the designated egfp concatenates. Although the Cre-mediated recombination is site specific, it is dependent on a randomly placed target site. That is, a genomic target cannot be preselected for integration based solely on its sequence. Conclusively, an rfp reporter gene was successfully inserted into the egfp target locus of zebrafish genome by Cre-lox-mediated recombination. This site-directed knockin system using the lox71/lox66 combination should be a promising gene-targeting platform serving various purposes in fish genetic engineering.

Correcting the systematic error of the density functional theory calculation: the alternate combination approach of genetic algorithm and neural network

The alternate combinational approach of genetic algorithm and neural network (AGANN) has been presented to correct the systematic error of the density functional theory (DFT) calculation. It treats the DFT as a black box and models the error through external statistical information. As a demonstration, the AGANN method has been applied in the correction of the lattice energies from the DFT calculation for 72 metal halides and hydrides. Through the AGANN correction, the mean absolute value of the relative errors of the calculated lattice energies to the experimental values decreases from 4.93% to 1.20% in the testing set. For comparison, the neural network approach reduces the mean value to 2.56%. And for the common combinational approach of genetic algorithm and neural network, the value drops to 2.15%. The multiple linear regression method almost has no correction effect here.

Comparison and combination of several stress relief methods for cubic boron nitride films deposited by ion beam assisted deposition

Cubic boron nitride (c-BN) films were prepared by ion beam assisted deposition (IBAD) technique, and the stresses were primary estimated by measuring the frequency shifts in the infrared-absorption peaks of c-BN samples. To test the possible effects of other factors, dependencies of the c-BN transversal optical mode position on film thickness and c-BN content were investigated. Several methods for reducing the stress of c-BN films including annealing, high temperature deposition, two-stage process, and the addition of a small amount of Si were studied, in which the c-BN films with similar thickness and cubic phase content were used to evaluate the effects of the various stress relief methods. It was shown that all the methods can reduce the stress in c-BN films to various extents. Especially, the incorporation of a small amount of Si (2.3 at.%) can result in a remarkable stress relief from 8.4 to similar to 3.6 GPa whereas the c-BN content is nearly unaffected, although a slight degradation of the c-BN crystallinity is observed. The stress can be further reduced down below I GPa by combination of the addition of Si with the two-stage deposition process. (c) 2008 Elsevier B.V. All rights reserved.

Optical characteristics of InAs quantum dots capped with short period GaAs/InAs superlattices and InGaAs combination layers

Long wavelength light emission was realized by capping InAs quantum dots (QDs) with short period GaAs/InAs superlattices (SLs) and an InGaAs strain-reducing layer (SRL). The optical properties were systematically investigated by photoluminescence tests. With increasing the periods of SLs, the emission wavelength of InAs QDs shifts from 1.27 to 1.53 mum. We explain the redshift as a result of the increased QD height with the SLs and the reduced strain in the dot caused by InGaAs SRL. (C) 2004 Published by Elsevier Ltd.

Luminescence properties of self-assembled InAs/GaAs quantum dots covered by InAlAs and InGaAs combination strain-reducing layer

We report the photoluminescence (PL) and structural properties of self-assembled InAs/GaAs quantum dots (QDs) covered by In0.2Al0.8As and In0.2Ga0.8As combination strain-reducing layer (SRL). By introducing a thin InAlAs layer, the ground state emission wavelength redshifts, and the energy splitting between the ground and first-excited states increases to 85 meV at 10 K. The energy splitting further increases to 92 meV and the temperature dependence of full width at half maximum (FWHM) changes for QDs with different SRL after the multi-stacking. These results are attributed to the fact that the combination layer has different effects on QDs compared to the InGaAs SRL.

Effect of In0.2Ga0.8As and In0.2Al0.8As combination layer on band offsets of InAs quantum dots

We demonstrate the self-organized InAs quantum dots capped with thin and In0.2Al0.8As and In0.2Ga0.8As combination layers with a large ground and first excited energy separation emission at 1.35 mum at room temperature. Deep level transient spectroscopy is used to obtain quantitative information on emission activation energies and capture barriers for electrons and holes. For this system, the emission activation energies are larger than those for InAs/GaAs quantum dots. With the properties of wide energy separation and deep emission activation energies, self-organized InAs quantum dots capped with In0.2Al0.8As and In0.2Ga0.8As combination layers are one of the promising epitaxial structures of 1.3 mum quantum dot devices. (C) 2004 American Institute of Physics.