Gene transfer techniques

Gene transfer techniques Success of gene therapy lies in efficient gene transfer into the cell. The gene (cDNA) is generally cloned into a vector to be able to deposit the foreign gene into the target cell. Selection of the right vector is crucial to gene ther-apy. An ideal vector should be able to protect and deliver DNA easily across the cell membrane into the nucleus, should have the ability to regulate expression of the gene of interest and minimize toxicity by targeting gene delivery to specific cells. It should be easy and inexpensive to produce in large quantities. Once the therapeutic gene is cloned into a vector with appropriate regulatory sequences (promoter/enhancer), it is introduced into the target cells. The genes can be delivered either ex vivo –where cells from a selected tissue of the patient are removed, exposed to the gene-transfer vector, selected forthe transgene using markers, and then the genetically cor-rected cells are reintroduced into the patient’s body; or in vivo where the vector DNA is injected directly into the body, generally into the tissue to be treated. Physical and chemical methods of gene transfer Various methods have evolved in the past few years to transfer genes to the target cells. Physical methods such as (a) microinjection of DNA into the cells or (b) electroporation, although very efficient, have their drawbacks in delivering genes in vivo. Also they are expensive as they involve use of specialized instruments. Chemical methods such as (a) calcium phosphate precipitation, where DNA in trapped in a fine precipitate which is endocytosed by the cell, or (b) DNA bound to the positively charged molecules such as DEAE-dextran or polybrene which then bind to the negatively charged cell membrane, are commonly used in the laboratories. DNA encapsulated in synthetic cationic lipid vesicles which fuse with the cell membrane and release DNA into the cell are being used in a number of gene therapy trials 2. Cationic liposome-mediated gene transfer is a safe and effective means of delivering genes directly into tumours. This approach prevents undesirable side effects.

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Gene Therapy for Cancer

Gene therapy, the latest development in the field of medicine, is based on manipulating, or changing the genetic material to treat and prevent the appearance of certain diseases that are difficult to heal with normal treatment. Gene therapy involves the introduction of genes, the biological unit of heredity, in cells or tissues of people who suffer from diseases like cancer. Gene therapy is done for the first time on September 14, 1990 by a group of physicians, consisting of W. French Anderson, MD and his colleagues Michael R. Blaese, MD, Bouza C. Institute and Kenneth Culver, MD, U. S. National Health. Performed in a four-year-old child suffering from "severe immunodeficiency combined, a rare genetic disease. Gene therapy can be classified into two main types of germ-line gene therapy and somatic gene therapy. Germline gene therapy involves the introduction of functional genes in the germ or reproductive cells (sperm and eggs) from the body, while in the case of somatic gene therapy, therapeutic genes are introduced into somatic cells. Gene therapy consists primarily in changing the genetic material (DNA and genes), and plays a key role in determining individual traits and characteristics. It has added a new dimension to cancer treatment, which is caused by mutation or DNA damage that leads to uncontrolled growth of abnormal cells. Most research was conducted and some are still underway to discover the potential of gene therapy in the treatment of breast, lung, prostate, bone, and leukemia. Different approaches for gene therapy to treat cancer, mainly concentrated or destroy or prevent growth of cancer cells or improve the ability of normal cells to fight cancer cells. If cancer is caused due to missing or changed genes, then gene therapy is the replacement of these genes with healthy ones. In addition, gene therapy can be done to stimulate the immune system to attack cancer cells. Through this technology, genes can be inserted into the patient's body, which, to instruct the cancer cells to produce certain proteins to inhibit stimulated Oncogenes cause cancer or tumor suppressor genes. Several other studies also underway to introduce genes into cancer cells, which can help make cancer cells more sensitive to various cancer treatments, including chemotherapy and radiotherapy. In addition, investigations are also underway to reduce the side effects of anticancer drugs to different cells increased resistance to flow. However, gene therapy, genes are introduced directly into the patient's body, but the virus used for this purpose. The virus is generally used for this therapy are retrovirus, adenovirus, herpes virus, lentivirus and poxviruses. Sometimes, liposomes (a small vesicle in a cell to shops and transport of substances in a cell) is also used as carriers in gene therapy. The virus can be used for both ex vivo and in vivo gene therapy. Ex vivo gene therapy involves the collection of blood or less bone marrow cells of the patient. Then the virus to insert genes into cells are needed in a laboratory, then injected into the patient's body. Furthermore, in vivo gene therapy involves the direct introduction of viruses or liposomes containing the desired gene in the patient's body. However, gene therapy is not without its drawbacks. One of the potential risks associated with gene therapy on the possibility of infection of healthy cells caused by the virus used to deliver the gene. Moreover, if genetic material is accidentally introduced into germ cells, then changes induced by it, will become the next generation. Again, this is very important to insert the desired gene in the right place, which can lead to failure and cause genetic mutations and cancer. More research is needed to eliminate the shortcomings of gene therapy, so you can truly revolutionize the treatment of life-threatening diseases like cancer.

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Gene doping

Gene doping is defined by the World Anti-Doping Agency as "the non-therapeutic use of cells, genes, genetic elements, or of the modulation of gene expression, having the capacity to improve athletic performance". A complex ethical and philosophical issue is what defines "gene doping", especially in the context of bioethical debates about human enhancement. An example of gene doping could involve the recreational use of gene therapies intended to treat muscle-wasting disorders. Many of these chemicals may be indistinguishable from their natural counterparts. In such cases, nothing unusual would enter the bloodstream so officials would detect nothing in a blood or urine test.

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Genome mapping

Genome mapping is the creation of a genetic map assigning DNA fragments to chromosomes. When a genome is first investigated, this map is nonexistent. The map improves with the scientific progress and is perfect when the genomic DNA sequencing of the species has been completed. During this process, and for the investigation of differences in strain, the fragments are identified by small tags. These may be genetic markers (PCR products) or the unique sequence-dependent pattern of DNA-cutting enzymes. The ordering is derived from genetic observations (recombinant frequency) for these markers or in the second case from a computational integration of the fingerprinting data. The term "mapping" is used in two different but related contexts. Two different ways of mapping are distinguished. Genetic mapping uses classical genetic techniques (e.g. pedigree analysis or breeding experiments) to determine sequence features within a genome. Using modern molecular biology techniques for the same purpose is usually referred to as physical mapping.

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Types of Gene Therapy

Virtually all cells in the human body contain genes, making them potential targets for gene therapy. However, these cells can be divided into two major categories: somatic cells (most cells of the body) or cells of the germline (eggs or sperm). In theory it is possible to transform either somatic cells or germ cells. Gene therapy using germ line cells results in permanent changes that are passed down to subsequent generations. If done early in embryologic development, such as during preimplantation diagnosis and in vitro fertilization, the gene transfer could also occur in all cells of the developing embryo. The appeal of germ line gene therapy is its potential for offering a permanent therapeutic effect for all who inherit the target gene. Successful germ line therapies introduce the possibility of eliminating some diseases from a particular family, and ultimately from the population, forever. However, this also raises controversy. Some people view this type of therapy as unnatural, and liken it to "playing God." Others have concerns about the technical aspects. They worry that the genetic change propagated by germ line gene therapy may actually be deleterious and harmful, with the potential for unforeseen negative effects on future generations. Somatic cells are nonreproductive. Somatic cell therapy is viewed as a more conservative, safer approach because it affects only the targeted cells in the patient, and is not passed on to future generations. In other words, the therapeutic effect ends with the individual who receives the therapy. However, this type of therapy presents unique problems of its own. Often the effects of somatic cell therapy are short-lived. Because the cells of most tissues ultimately die and are replaced by new cells, repeated treatments over the course of the individual's life span are required to maintain the therapeutic effect. Transporting the gene to the target cells or tissue is also problematic. Regardless of these difficulties, however, somatic cell gene therapy is appropriate and acceptable for many disorders, including cystic fibrosis, muscular dystrophy, cancer, and certain infectious diseases. Clinicians can even perform this therapy in utero, potentially correcting or treating a life-threatening disorder that may significantly impair a baby's health or development if not treated before birth.

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Types of Gene Therapy

Gene therapy is the insertion of genes into an individual's cells and tissues to treat a disease, and hereditary diseases in which a defective mutant allele is replaced with a functional one. Although the technology is still in its infancy, it has been used with some success. Antisense therapy is not strictly a form of gene therapy, but is a genetically-mediated therapy and is often considered together with other methods. How does gene therapy work? In most gene therapy studies, a "normal" gene is inserted into the genome to replace an "abnormal," disease-causing gene. A carrier molecule called a vector must be used to deliver the therapeutic gene to the patient's target cells. Currently, the most common vector is a virus that has been genetically altered to carry normal human DNA. Viruses have evolved a way of encapsulating and delivering their genes to human cells in a pathogenic manner. Scientists have tried to take advantage of this capability and manipulate the virus genome to remove disease-causing genes and insert therapeutic genes. Target cells such as the patient's liver or lung cells are infected with the viral vector. The vector then unloads its genetic material containing the therapeutic human gene into the target cell. The generation of a functional protein product from the therapeutic gene restores the target cell to a normal state. How is gene therapy being studied in the treatment of cancer? Researchers are studying several ways to treat cancer using gene therapy. Some approaches target healthy cells to enhance their ability to fight cancer. Other approaches target cancer cells, to destroy them or prevent their growth. Some gene therapy techniques under study are described below. In one approach, researchers replace missing or altered genes with healthy genes. The types of gene therapy described thus far all have one factor in common: that is, that the tissues being treated are somatic (somatic cells include all the cells of the body, excluding sperm cells and egg cells). In contrast to this is the replacement of defective genes in the germline cells (which contribute to the genetic heritage of the offspring). Gene therapy in germline cells has the potential to affect not only the individual being treated, but also his or her children as well. Germline therapy would change the genetic pool of the entire human species, and future generations would have to live with that change. Gene Therapy: requirements The gene must be identified and cloned. This has been done for the ADA gene.It must be inserted in cells that can take up long-term residence in the patient. So far, this means removing the patient's own cells, treating them in tissue culture, and then returning them to the patient. It must be inserted in the DNA so that it will be expressed adequately; that is, transcribed and translated with sufficient efficiency that worthwhile amounts of the enzyme are produced. All these requirements seem to have been met for SCID therapy using a retrovirus as the gene vector. Retroviruses have several advantages for introducing genes into human cells. Their envelope protein enables the virus to infect human cells. RNA copies of the human ADA gene can be incorporated into the retroviral genome using a packaging cell.

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TYPES OF GENE THERAPY: -

Gene therapy may be classified into two types 1) Germ line gene therapy 2) Somatic cell gene therapy a) Incase of germ line gene therapy germ cells that is sperms or eggs are modified by the introduction of functional genes, which are ordinarily integrated into their genomes. Therefore the change due to therapy is heritable and passed onto the later generations. This approach, heretically, is highly effective in counteracting the genetic disorders. However this option is not consider, at least for the present for application in human beings for a variety of technical and ethical reasons. b) In the case of somatic cell gene therapy the gene is introduced only in somatic cells, especially of those tissues in which expression of the concerned gene is critical for health. Expression of the introduced gene relieves symptoms of the disorder, but this effect is not heritable, as it does not involve the germ line. It is the only feasible option, and clinical trials have already started mostly for the treatment of cancer and blood disorders. GENERAL GENE THERAPY STRATEGIES 1) Gene augmentation therapy (GAT): - It is done by simple addition of functional alleles has been used to treat several inherited disorders caused by genetic deficiency of a gene product. It is also involved in transfer to cells of genes encoding toxic compounds (suicide genes) or prodrugs (reagents which confer sensitivity to subsequent treatment with a drug). It has been particularly applied to autosomal recessive disorders where even modest expression levels of an introduced gene may make a substantial difference. 2) Targeted killing of specific cells: - Artificial cell killing and immune system assisted cell killing have been popular in the treatment of cancers. It can be done by two ways. a) Direct cell killing: - it is possible if the inserted genes are expressed to produce a lethal toxin (suicide genes), or a gene encoding a prodrug is inserted, conferring susceptibility to killing by a subsequently administered drug. Alternatively selectively lytic viruses can be used. b) Indirect cell killing: - It uses immunostimulatory genes to provoke or enhance an immune response against the target cell. 3) Targeted mutation correction: - The repair of a genetic defect to restore a functional allele, is the exception, technical difficulties have meant that it is not sufficiently reliable to warrant clinical trails. 4) Targeted inhibition of gene expression: - It is suitable for treating infectious diseases and some cancers. If disease cells display a novel gene product or inappropriate expression of a gene a variety of different systems can be used specifically to block the expression of a single gene at the DNA, RNA or Protein levels.

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Gene Therapy

The term gene therapy can be defined as introduction of a normal functional gene into cells, which contain the defective allele of concerned gene with the objective of correcting a genetic disorder or an acquired disorder. The first approach in gene therapy is: - a) Identification of the gene that plays the key role in the development of a genetic disorder. b) Determination of the role of its product in health and disease. c) Isolation and cloning of the gene. d) Development of an approach for gene therapy. The genetic material may be transferred directly into cells within a patient, which is referred as in vivo gene therapy or else cells may be removed from the patient and the genetic material inserted into them, which is referred as invitro gene therapy. Apart from the two methods mentioned above there is one more method that is ex-vivo gene therapy in which genetic material is inserted into the cells just prior to transplanting the modified cells back into the patient. Major disease classes under gene therapy include: - a) Infectious diseases: - infection by a virus or bacterial pathogen b) Cancers: - uncontrolled and enormous cell division and cell proliferation as a result of activation of an oncogene or inactivation of a tumors suppressor gene or an apoptosis gene. c) Inherited disorders: - genetic deficiency of an individual gene product or genetically determined in appropriate expression of a gene. d) Immune system disorders: - includes allergies, inflammation and also autoimmune diseases in which immune system cells appropriately destroy body cells.

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Cell division

Cell division is the process by which a parent cell divides into two or more daughter cells. Cell division is usually a small segment of a larger cell cycle. This type of cell division in eukaryotes is known as mitosis, and leaves the daughter cell capable of dividing again. The corresponding sort of cell division in prokaryotes is known as binary fission. In another type of cell division present only in eukaryotes, called meiosis, a cell is permanently transformed into a gamete and cannot divide again until fertilization. For simple unicellular organisms such as the amoeba, one cell division is equivalent to reproduction-- an entire new organism is created. On a larger scale, mitotic cell division can create progeny from multicellular organisms, such as plants that grow from cuttings. Cell division also enables asexually reproducing organisms to develop from the one-celled zygote, which itself was produced by cell division from gametes. And after growth, cell division allows for continual construction and repair of the organism. A human being's body experiences about 10,000 trillion cell divisions in a lifetime. The primary concern of cell division is the maintenance of the original cell's genome. Before division can occur, the genomic information which is stored in chromosomes must be replicated, and the duplicated genome separated cleanly between cells. A great deal of cellular infrastructure is involved in keeping genomic information consistent between "generations".

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Cell biology

Cell biology (formerly cytology, from the Greek kytos, "container") is an academic discipline that studies cells – their physiological properties, their structure, the organelles they contain, interactions with their environment, their life cycle, division and death. This is done both on a microscopic and molecular level. Cell biology research encompasses both the great diversity of single-celled organisms like bacteria and protozoa, as well as the many specialized cells in multicellular organisms like humans. Knowing the components of cells and how cells work is fundamental to all biological sciences. Appreciating the similarities and differences between cell types is particularly important to the fields of cell and molecular biology as well as to biomedical fields such as cancer research and developmental biology. These fundamental similarities and differences provide a unifying theme, sometimes allowing the principles learned from studying one cell type to be extrapolated and generalized to other cell types. Hence, research in cell biology is closely related to genetics, biochemistry, molecular biology, Immunology, and developmental biology.

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Happy New Year 2010

Wish You All Happy New Year. Please Welcome the new year 2010

We are post about the Nucleic Acids - DNA, RNA -Genetics .all sources are taken from the internet we didn’t create anything or discover thing.Just we collect for you all. Thanks For visiting our blog www.biomolecules-world.blogspot.com/.Thanks for supporting us. Please support us for this coming year also. DNA,GENE,DNA VS RNA,Bio Molecules,DNA Testing centers,DNA Implementation,RNA Implementation,Genetic testing,DNA sequencing

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