No abstract available.
Transposases*

Transposons are the mobile and autonomic replication DNA fragments in genomes. With more understanding of the structure and function of transposons, numerous transposons have been developed to the genetics tool for gene function analysis, gene transformation and gene therapy. The low transpositional activity of the natural transposons is the main obstacles to the utilization of transposons. Recently, with the progress in bioinformatics and protein engineering methods, researchers have reconstructed and optimized natural transposases to create hyperactive transposases that catalyze the transposition with high efficiency. The resulted hyperactive transposons have been applied to gene-modification and gene-tagging. Meanwhile, transposase chimeras were created by protein fusion technology. The insertion characteristic of transposons were artificially regulated which could be utilized in gene therapy.
DNA Transposable Elements ; Gene Targeting ; Genetic Therapy ; Protein Engineering ; Transposases ; chemistry

PiggyBac is a transposable DNA element originally discovered in the cabbage looper moth (Trichoplusia ni). The T. ni piggyBac transposon can introduce exogenous fragments into a genome, constructing a transgenic organism. Nevertheless, the comprehensive analysis of endogenous piggyBac-like elements (PLEs) is important before using piggyBac, because they may influence the genetic stability of transgenic lines. Herein, we conducted a genome-wide analysis of PLEs in the brown planthopper (BPH) Nilaparvata lugens (Stål) (Hemiptera: Delphacidae), and identified a total of 28 PLE sequences. All N. lugens piggyBac-like elements (NlPLEs) were present as multiple copies in the genome of BPH. Among the identified NlPLEs, NlPLE25 had the highest copy number and it was distributed on five chromosomes. The full length of NlPLE25 consisted of terminal inverted repeats and sub-terminal inverted repeats at both terminals, as well as a single open reading frame transposase encoding 546 amino acids. Furthermore, NlPLE25 transposase caused precise excision and transposition in cultured insect cells and also restored the original TTAA target sequence after excision. A cross-recognition between the NlPLE25 transposon and the piggyBac transposon was also revealed in this study. These findings provide useful information for the construction of transgenic insect lines.
Amino Acid Sequence ; Animals ; Animals, Genetically Modified ; DNA Transposable Elements/genetics* ; Hemiptera/genetics* ; Transposases/genetics*

Despite recent economic prosperity, Korea still has high prevalence of tuberculosis. Molecular biologic characterization of Korean Mycobacterium tuberculosis strains might provide a deeper understanding of the forces contributing to the spread of tuberculosis in Korea. Therefore, we analyzed the cell lysate proteome of a representative Korean Mycobacterium tuberculosis isolate (K01) in comparison with laboratory reference strains H37Rv and H37Ra. Seven spots were strongly expressed only in K01 strain compared with M. tuberculosis H37Rv and H37Ra. Through continuous MALDI-MS analysis, these spots were identified as hypothetical protein Rv3849, secreted immunogenic protein Mpt64, Acetyl/propionyl-CoA Carbpxylase (AccD1), alkyl hydroperoxide reductase C (AhpC), N-acetylmuramyl-L-alanine amidase, a putative UDP glucose epimerase, and a transposase. A deeper study of these proteins may provide a clue in the development of effective new anti-tuberculosis vaccines against Korean M. tuberculosis isolates.
Korea ; Mycobacterium tuberculosis* ; Mycobacterium* ; Peroxiredoxins ; Prevalence ; Proteome* ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Transposases ; Tuberculosis ; UDPglucose 4-Epimerase ; Vaccines

No abstract available.
Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/*genetics ; Carrier Proteins/*genetics ; Drug Resistance, Multiple, Bacterial/*genetics ; Humans ; Stenotrophomonas maltophilia/drug effects/*genetics/isolation & purification ; Transposases/*genetics ; Trimethoprim, Sulfamethoxazole Drug Combination/*pharmacology

In an effort to generate a desired expression construct for making heart-specific expression transgenic zebrafish, a Tol2 plasmid, which can drive EGFP reporter gene specifically expressed in the heart, was modified using subcloning technology. An IRES fragment bearing multiple cloning site (MCS) was amplified directly from pIRES2-EGFP plasmid and was inserted between the CMLC2 promoter and EGFP fragment of the pDestTol2CG vector. This recombinant expression plasmid pTol2-CMLC2-IRES-EGFP can drive any interested gene specifically expressed in the zebrafish heart along with EGFP reporter gene. To test the effectiveness of this new expression plasmid, we constructed pTol2-CMLC2-RED-IRES-EGFP plasmid by inserting another reporter gene DsRed-Monome into MCS downstream of the CMLC2 promoter and injected this transgenic recombinant plasmid into one-cell stage embryos of zebrafish. Under fluorescence microscope, both the red fluorescence and the green fluorescence produced by pTol2-CMLC2-RED-IRES-EGFP were detected specifically in the heart tissue in the same expression pattern. This novel expression construct pTol2-CMLC2-IRES-EGFP will become an important tool for our research on identifying heart development candidate genes' function using zebrafish as a model.
Animals ; Animals, Genetically Modified ; genetics ; growth & development ; DNA Transposable Elements ; genetics ; Genes, Reporter ; genetics ; Genetic Vectors ; biosynthesis ; genetics ; Green Fluorescent Proteins ; genetics ; Myocardium ; metabolism ; Plasmids ; genetics ; Transfection ; Transgenes ; Transposases ; genetics ; Zebrafish ; genetics ; Zebrafish Proteins ; genetics ; metabolism