Progress in gene transfer by germ cells in mammals

Use of germ cells as vectors for transgenesis in mammals has been well developed and offers exciting prospects for experimental and applied biology, agricultural and medical sciences.Such approach is referred to as either male germ cell mediated gene transfer (MGCMGT)or female germ cell mediated gene transfer(FGCMGT)technique.Sperm-mediated gene transfer (SMGT),including its alternative method,testis-mediated gene transfer(TMGT),becomes an established and reliable method for transgenesis.They have been extensively used for producing transgenic animals.The newly developed approach of FGCMGT,ovary-mediated gene transfer(OMGT) is also a novel and useful tool for efficient transgenesis.This review highlights an overview of the recent progress in germ cell mediated gene transfer techniques,methods developed and mechanisms of nucleic acid uptake by germ cells.     

基因治疗心肌缺血的载体应用新进展

近年来,随着基因治疗技术的不断进步,为心肌缺血的治疗开辟了一条全新的途径,并取得了一些令人鼓舞的进展。基因治疗主要包括治疗基因、基因转移载体以及基因导入途径三个方面。基因转移载体又在治疗基因和基因表达之间起着桥梁作用,因此,发展安全、高效的基因转移系统是基因治疗的关键之一。目前用于基因治疗心肌缺血基因转移的载体主要有病毒载体和非病毒载体。下面将就不同载体在心肌缺血的基因治疗中的应用进展进行简要的总结。     

基因治疗心肌缺血的载体应用新进展

近年来,随着基因治疗技术的不断进步,为心肌缺血的治疗开辟了一条全新的途径,并取得了一些令人鼓舞的进展。基因治疗主要包括治疗基因、基因转移载体以及基因导入途径三个方面。基因转移载体又在治疗基因和基因表达之间起着桥梁作用,因此,发展安全、高效的基因转移系统是基因治疗的关键之一。目前用于基因治疗心肌缺血基因转移的载体主要有病毒载体和非病毒载体。下面将就不同载体在心肌缺血的基因治疗中的应用进展进行简要的总结。     

Techniques for gene transfer into neurons

To illuminate the function of the thousands of genes that make up the complexity of the nervous system, it is critical to be able to introduce and express DNA in neurons. Over the past two decades, many gene transfer methods have been developed, including viral vectors, liposomes and electroporation. Although the perfect gene transfer technique for every application has not yet been developed, recent technical advances have facilitated the ease of neuronal gene transfer and have increased the accessibility of these techniques to all laboratories. In order to select a transfection method for any particular experiment, the specific advantages and disadvantages of each technique must be considered.     

Relations between animal transgenesis and reproduction

Transgenesis has become an essential tool for the study of gene expression mechanisms and functions. Transgenesis is also more and more used for biotechnological applications such as the study of human diseases, the adaptation of pig organs to humans, the production of pharmaceutical proteins in milk and likely in the future for the improvement of animal production. The use of transgenesis relies on the efficiency of gene transfer. New tools have been recently designed to improve gene transfer. The methods of gene transfer are highly dependent on the techniques of animal reproduction. Conversely, the need to improve transgenesis urges researchers to study some of the key steps in reproduction and to find new techniques for gene transfer. This paper summarises the recent data and the perspectives offered by animal transgenesis.     

Nonviral gene therapy targeting cardiovascular system

The goal of gene therapy is either to introduce a therapeutic gene into or replace a defective gene in an individual's cells and tissues. Gene therapy has been urged as a potential method to induce therapeutic angiogenesis in ischemic myocardium and peripheral tissues after extensive investigation in recent preclinical and clinical studies. A successful gene therapy mainly relies on the development of the gene delivery vector. Developments in viral and nonviral vector technology including cell-based gene transfer will further improve transgene delivery and expression efficiency. Nonviral approaches as alternative gene delivery vehicles to viral vectors have received significant attention. Recently, a simple and safe approach of gene delivery into target cells using naked DNA has been improved by combining several techniques. Among the physical approaches, ultrasonic microbubble gene delivery, with its high safety profile, low costs, and repeatable applicability, can increase the permeability of cell membrane to macromolecules such as plasmid DNA by its bioeffects and can provide as a feasible tool in gene delivery. On the other hand, among the promising areas for gene therapy in acquired diseases, ischemic cardiovascular diseases have been widely studied. As a result, gene therapy using advanced technology may play an important role in this regard. The aims of this review focus on understanding the cellular and in vivo barriers in gene transfer and provide an overview of currently used chemical vectors and physical tools that are applied in nonviral cardiovascular gene transfer.     

Genetic engineering in marine cyanobacteria

Many species of microalgae producing useful materials have been isolated from marine environments. For their industrial application, widely applicable and stable gene expression is required. It is necessary to establish gene transfer methods as an essential first step in genetic manipulation. Although gene transfer techniques for cyanobacteria have been developed, only naturally transformable strains have been used. Here, we describe recent progress made in developing gene transfer methods for marine cyanobacteria. The following are covered: (1) transformation, (2) electroporation, (3) conjugation, (4) particle gun. A plasmid from the marine cyanobacterium, Synechococcus sp., whose copy number is dependent on salinity, was characterized. This plasmid is being used to develop a stable and controllable gene expression system.     

DNA-mediated genetic transformation of mouse embryos and bone marrow--a review

In recent years, new gene transfer systems have been developed which allow molecularly cloned genetic material to be introduced into whole organisms. These systems include the microinjection of DNA into mammalian embryos, transfection of DNA into mouse bone marrow cells, and the infection of early embryos with retroviruses. Exogenous DNA appears to integrate randomly into the host genome. The production of transgenic mice by injection of DNA into mouse embryos has rapidly gained importance as an experimental tool for the study of gene regulation during development. Through this technique, recombinant molecules of any type can be introduced into one-celled embryos, and thus can be used to study development from its earliest stages. DNA sequences have been shown to integrate and transmit through the germ line to subsequent generations as mendelian traits. Transgenic mice carrying various gene constructs have been successfully exploited for the elucidation of factors which determine tissue specificity of gene expression as well as the level of gene control. Phenotypic changes related to expression of foreign genes have also been observed. This experimental approach thus promises to rapidly solve many of the heretofore most challenging problems in developmental genetics. Insertion of foreign genes has also made possible the creation of insertional mutants which manifest themselves most frequently as recessives. Such mutations can be readily studied at the molecular level by using the transferred material as a probe for recovery of the affected host sequence from genomic libraries. Many of these same problems have been addressed by introducing retroviral DNA into mouse embryos. Here, the sequences used for transfer have been limited to retroviral genes, but nonetheless these experiments have been profitably exploited for studies both of gene regulation and mutagenesis. Gene transfer systems are being developed allowing the experimenter to transfer DNA into bone marrow cells of mice, after which the recipient cells can be reintroduced into lethally irradiated histocompatible animals. This system has the advantage that selection can be applied during the gene transfer process such that the expression of the foreign material is assured. In addition, these experiments have created a model system for production of animals carrying a subpopulation of cells which is highly resistant to a toxic agent. This system has the potential for therapeutic application to man.     

About the last common ancestor, the universal life-tree and lateral gene transfer: a reappraisal

An organismal tree rooted in the bacterial branch and derived from a hyperthermophilic last common ancestor (LCA) is still widely assumed to represent the path followed by evolution from the most primeval cells to the three domains recognized among contemporary organisms: Bacteria, Archaea and Eucarya. In the past few years, however, more and more discrepancies between this pattern and individual protein trees have been brought to light. There has been an overall tendency to attribute these incongruities to widespread lateral gene transfer. However, recent developments, a reappraisal of earlier evidence and considerations of our own lead us to a quite different view. It would appear (i) that the role of lateral gene transfer was overemphasized in recent discussions of molecular phylogenies; (ii) that the LCA was probably a non-thermophilic protoeukaryote from which both Archaea and Bacteria emerged by reductive evolution but not as sister groups, in keeping with a current evolutionary scheme for the biosynthesis of membrane lipids; and (iii) that thermophilic Archaea may have been the first branch to diverge from the ancestral line.     

Evolutionary analyses of the small subunit of glutamate synthase: gene order conservation,gene fusions,and prokaryote-to-eukaryote lateral gene transfers

Lateral gene transfer has been identified as an important mode of genome evolution within prokaryotes. Except for the special case of gene transfer from organelle genomes to the eukaryotic nucleus, only a few cases of lateral gene transfer involving eukaryotes have been described. Here we present phylogenetic and gene order analyses on the small subunit of glutamate synthase (encoded by gltD) and its homologues, including the large subunit of sulfide dehydrogenase (encoded by sudA). The scattered distribution of the sudA and sudB gene pair and the phylogenetic analysis strongly suggest that lateral gene transfer was involved in the propagation of the genes in the three domains of life. One of these transfers most likely occurred between a prokaryote and an ancestor of diplomonad protists. Furthermore, phylogenetic analyses indicate that the gene for the small subunit of glutamate synthase was transferred from a low-GC gram-positive bacterium to a common ancestor of animals, fungi, and plants. Interestingly, in both examples, the eukaryotes encode a single gene that corresponds to a conserved operon structure in prokaryotes. Our analyses, together with several recent publications, show that lateral gene transfers from prokaryotes to unicellular eukaryotes occur with appreciable frequency. In the case of the genes for sulfide dehydrogenase, the transfer affected only a limited group of eukaryotes—the diplomonads—while the transfer of the glutamate synthase gene probably happened earlier in evolution and affected a wider range of eukaryotes.     

Intergeneric transfer of ribosomal genes between two fungi

Background  

Horizontal gene transfer, also called lateral gene transfer, frequently occurs among prokaryotic organisms, and is considered an important force in their evolution. However, there are relatively few reports of transfer to or from fungi, with some notable exceptions in the acquisition of prokaryotic genes. Some fungal species have been found to contain sequences resembling those of bacterial genes, and with such sequences absent in other fungal species, this has been interpreted as horizontal gene transfer. Similarly, a few fungi have been found to contain genes absent in close relatives but present in more distantly related taxa, and horizontal gene transfer has been invoked as a parsimonious explanation. There is a paucity of direct experimental evidence demonstrating the occurrence of horizontal gene transfer in fungi.     

Gene duplication,transfer, and evolution in the chloroplast genome

In addition to the nuclear genome, organisms have organelle genomes. Most of the DNA present in eukaryotic organisms is located in the cell nucleus. Chloroplasts have independent genomes which are inherited from the mother. Duplicated genes are common in the genomes of all organisms. It is believed that gene duplication is the most important step for the origin of genetic variation, leading to the creation of new genes and new gene functions. Despite the fact that extensive gene duplications are rare among the chloroplast genome, gene duplication in the chloroplast genome is an essential source of new genetic functions and a mechanism of neo-evolution. The events of gene transfer between the chloroplast genome and nuclear genome via duplication and subsequent recombination are important processes in evolution. The duplicated gene or genome in the nucleus has been the subject of several recent reviews. In this review, we will briefly summarize gene duplication and evolution in the chloroplast genome. Also, we will provide an overview of gene transfer events between chloroplast and nuclear genomes.     

Ultrasound: mechanical gene transfer into plant cells by sonoporation

Development of nonviral gene transfer methods would be a valuable alternative of gene therapy or transformation. Ultrasound can produce a variety of nonthermal bioeffects via acoustic cavitation. Cavitation bubbles can induce cell death or transient membrane permeabilization (sonoporation) on cells. Application of sonoporation for gene transfer into cells or tissues develops quickly in recent years. Many studies have been performed in vitro exposure systems to a variety of cell lines transfected successfully. In vivo, cavitation initiation and control are more difficult, but can be enhanced by ultrasound contrast agents (microbubbles). The use of ultrasound for nonviral gene delivery has been applied for mammalian systems, which provides a fundamental basis and strong promise for development of new gene therapy methods for clinical medicine. In this paper, ultrasound applied to plant cell transformation or gene transfer is reviewed. Recently, most researches are focused on sonication-assisted Agrobacterium-mediated transformation (SAAT) in plant cells or tissues. Microbubbles are also proposed to apply to gene transfer in plant cells and tissues.     

Computational and phylogenetic validation of nematode horizontal gene transfer

Sequencing of expressed genes has shown that nematodes, particularly the plant-parasitic nematodes, have genes purportedly acquired from other kingdoms by horizontal gene transfer. The prevailing orthodoxy is that such transfer has been a driving force in the evolution of niche specificity, and a recent paper in BMC Evolutionary Biology that presents a detailed phylogenetic analysis of cellulase genes in the free-living nematode Pristionchus pacificus at the species, genus and family levels substantiates this hypothesis.     

Host-specific toxins: effectors of necrotrophic pathogenicity

Host-specific toxins (HSTs) are defined as pathogen effectors that induce toxicity and promote disease only in the host species and only in genotypes of that host expressing a specific and often dominant susceptibility gene. They are a feature of a small but well-studied group of fungal plant pathogens. Classical HST pathogens include species of Cochliobolus , Alternaria and Pyrenophora . Recent studies have shown that Stagonospora nodorum produces at least four separate HSTs that interact with four of the many quantitative resistance loci found in the host, wheat. Rationalization of fungal phylogenetics has placed these pathogens in the Pleosporales order of the class Dothideomycetes. It is possible that all HST pathogens lie in this order. Strong evidence of the recent lateral gene transfer of the ToxA gene from S. nodorum to Pyrenophora tritici-repentis has been obtained. Hallmarks of lateral gene transfer are present for all the studied HST genes although definitive proof is lacking. We therefore suggest that the Pleosporales pathogens may have a conserved propensity to acquire HST genes by lateral transfer.     

Gene therapy for muscle disease

The molecular mechanisms of Duchenne muscular dystrophy (DMD) have been extensively investigated since the discovery of the dystrophin gene in 1986. Nonetheless, there is currently no effective treatment for DMD. Recent reports, however, indicate that adenoassociated viral (AAV) vector-mediated transfer of a functional dystrophin cDNA into the affected muscle is a promising strategy. In addition, antisense-mediated exon skipping technology has been emerging as another promising approach to restore dystrophin expression in DMD muscle. Ongoing clinical trials show restoration of dystrophin in DMD patients without serious side effects. Here, we summarize the recent progress in gene therapy, with an emphasis on exon skipping for DMD.     

Horizontal gene transfer in eukaryotic evolution

Horizontal gene transfer (HGT; also known as lateral gene transfer) has had an important role in eukaryotic genome evolution, but its importance is often overshadowed by the greater prevalence and our more advanced understanding of gene transfer in prokaryotes. Recurrent endosymbioses and the generally poor sampling of most nuclear genes from diverse lineages have also complicated the search for transferred genes. Nevertheless, the number of well-supported cases of transfer from both prokaryotes and eukaryotes, many with significant functional implications, is now expanding rapidly. Major recent trends include the important role of HGT in adaptation to certain specialized niches and the highly variable impact of HGT in different lineages.     

Gene therapy techniques.

The most dramatic event of the past year in the field of gene therapy has been the initiation of clinical trials involving the introduction of genetically altered cells into human beings. Four studies, three involving new approaches to cancer therapy and one involving the treatment of adenosine deaminase deficiency, are presently under way. There has also been significant recent progress in the technology of gene transfer relevant to gene therapy. This progress, along with the recent clinical therapy trials, is the subject of this review.