DNA -BIOTECHNOLOGY PORTAL

DNA -BIOTECHNOLOGY PORTAL

VISIT http://www.indiareviewchannel.com FOR THE BEST REVIEWS FOR HEALTH PRODUCTS... DNA is composed of deoxyribonucleic acid.in the DNA the base sequence is of paramount importance.the
genetic information is coded in the specific sequence of bases.in the DNA the base sequence is always written
from the 5' end to the 3'end.this is called the polarity of the DNA chain..the DNA consists of two polydeoxyribonucleotide chains.
the DNA strands are held togethermainly by hydrogen bonds between purine and pyrimidine bases.the two DNA strands run antiparallel.
in a cell about 90% DNA are permanently inactive.histones and specific proteins help in this process of inactivation and subsequent
differentiation.



RECOMBINANT DNA TECHNOLOGY

genetic recombination is the exchange of information between two DNA segments. this is a common occurancewithin the same species.
but by artificial means when a gene of one species is transferred to another living organism,it is called RECOMBINANT DNA TECHNOLOGY. in
common parlance , this is known as GENETIC ENGINEERING.

DNA finger printing in forensic medicine

dna probes are developed so that at low stringency , they hybridise to anumber of these loci to produce individual specific finger prints.DNA profile is
used in this technique.

ISOTOPES

ISOTOPES are used in RADIO ACTIVITY.
ISOTOPES are the elements having the same atomic number but different mass number.the spontaneous degradation of nucleusand transmutation of one element to another with consequent emission
of rays or particles is known as RADIOACTIVITY.

Google Enter your search terms Submit search form

Oligonucleotides
The use of synthetic oligonucleotides in gene therapy is to inactivate the genes involved in the disease process. There are several methods by which this is achieved. One strategy uses antisense specific to the target gene to disrupt the transcription of the faulty gene. Another uses small catalytic molecules of RNA called siRNA to cleave specific unique sequences in the mRNA transcript of the faulty gene, disrupting translation of the faulty mRNA, and therefore expression of the gene. A further strategy uses double stranded oligodeoxynucleotides as a decoy for the transcription factors that are required to activate the transcription of the target gene. The transcription factors bind to the decoys instead of the promoter of the faulty gene, which reduces the transcription of the target gene, lowering expression.

Lipoplexes and Polyplexes
To improve the delivery of the new DNA into the cell, the DNA must be protected from damage and its entry into the cell must be facilitated. To this end new molecules, lipoplexes and polyplexes, have been created that have the ability to protect the DNA from undesirable degradation during the transfection process. Plasmid DNA can be covered with lipids in an organized structure like a micelle or a liposome. When the organized structure is complexed with DNA it is called a lipoplex. There are three types of lipids, anionic (negatively charged), neutral, or cationic (positively charged). Initially, anionic and neutral lipids were used for the construction of lipoplexes for synthetic vectors. However, in spite of the facts that there is little toxicity associated with them, that they are compatible with body fluids and that there was a possibility of adapting them to be tissue specific; they are complicated and time consuming to produce so attention was turned to the cationic versions. Cationic lipids, due to their positive charge, naturally complex with the negatively charged DNA. Also as a result of their charge they interact with the cell membrane, endocytosis of the lipoplex occurs and the DNA is released into the cytoplasm. The cationic lipids also protect against degradation of the DNA by the cell. The most common use of lipoplexes has been in gene transfer into cancer cells, where the supplied genes have activated tumor suppressor control genes in the cell and decrease the activity of oncogenes. Recent studies have shown lipoplexes to be useful in transfecting respiratory epithelial cells, so they may be used for treatment of genetic respiratory diseases such as cystic fibrosis. Complexes of polymers with DNA are called polyplexes. Most polyplexes consist of cationic polymers and their production is regulated by ionic interactions. One large difference between the methods of action of polyplexes and lipoplexes is that polyplexes cannot release their DNA load into the cytoplasm, so to this end, co-transfection with endosome-lytic agents (to lyse the endosome that is made during endocytosis, the process by which the polyplex enters the cell) such as inactivated adenovirus must occur. However this isn't always the case, polymers such as polyethylenimine have their own method of endosome disruption as does chitosan and trimethylchitosan.

Recent Developments In Gene Therapy




Scientists at the National Institutes of Health (Bethesda, Maryland) have successfully treated metastatic melanoma in two patients using killer T cells genetically retargeted to attack the cancer cells. This study constitutes the first demonstration that gene therapy can be effective in treating cancer. The study results have been published in Science (October 2006). In May 2006 a team of scientists led by Dr. Luigi Naldini and Dr. Brian Brown from the San Raffaele Telethon Institute for Gene Therapy (HSR-TIGET) in Milan, Italy reported a breakthrough for gene therapy in which they developed a way to prevent the immune system from rejecting a newly delivered gene. Similar to organ transplantation, gene therapy has been plagued by the problem of immune rejection. So far, delivery of the 'normal' gene has been difficult because the immune system does not recognize the new gene and rejects the cells carrying it. To overcome this problem, the HSR-TIGET group utilized a newly uncovered network of genes regulated by molecules known as microRNAs. Dr. Naldini's group reasoned that they could use this natural function of microRNA to selectively turn off the identity of their therapeutic gene in cells of the immune system and prevent the gene from being found and destroyed. The researchers injected mice with the gene containing an immune-cell microRNA target sequence, and spectacularly, the mice did not reject the gene, as previously occurred when vectors without the microRNA target sequence were used. This work will have important implications for the treatment of hemophilia and other genetic diseases by gene therapy. In March 2006 an international group of scientists announced the successful use of gene therapy to treat two adult patients for a disease affecting myeloid cells. The study, published in Nature Medicine, is believed to be the first to show that gene therapy can cure diseases of the myeloid system. In 2003 a University of California, Los Angeles research team inserted genes into the brain using liposomes coated in a polymer called polyethylene glycol (PEG). The transfer of genes into the brain is a significant achievement because viral vectors are too big to get across the "blood-brain barrier." This method has potential for treating Parkinson's disease. See Undercover genes slip into the brain at NewScientist.com (March 20, 2003). RNA interference or gene silencing may be a new way to treat Huntington's. Short pieces of double-stranded RNA (short, interfering RNAs or siRNAs) are used by cells to degrade RNA of a particular sequence. If a siRNA is designed to match the RNA copied from a faulty gene, then the abnormal protein product of that gene will not be produced. See Gene therapy may switch off Huntington's at NewScientist.com.

Envelope Protein Pseudotyping Of Viral Vectors
The viral vectors described above have natural host cell populations that they infect most efficiently. Retroviruses have limited natural host cell ranges, and although adenovirus and adeno-associated virus are able to infect a relatively broader range of cells efficiently, some cell types are refractory to infection by these viruses as well. Attachment to and entry into a susceptible cell is mediated by the protein envelope on the surface of a virus. Retroviruses and adeno-associated viruses have a single protein coating their membrane, while adenoviruses are coated with both an envelope protein and fibers that extend away from the surface of the virus. The envelope proteins on each of these viruses bind to cell-surface molecules such as heparin sulfate, which localizes them upon the surface of the potential host, as well as with the specific protein receptor that either induces entry-promoting structural changes in the viral protein, or localizes the virus in endosomes wherein acidification of the lumen induces this refolding of the viral coat. In either case, entry into potential host cells requires a favorable interaction between a protein on the surface of the virus and a protein on the surface of the cell. For the purposes of gene therapy, one might either want to limit or expand the range of cells susceptible to transduction by a gene therapy vector. To this end, many vectors have been developed in which the endogenous viral envelope proteins have been replaced by either envelope proteins from other viruses, or by chimeric proteins. Such chimera would consist of those parts of the viral protein necessary for incorporation into the virion as well as sequences meant to interact with specific host cell proteins. Viruses in which the envelope proteins have been replaced as described are referred to as pseudotyped viruses. For example, the most popular retroviral vector for use in gene therapy trials has been the lentivirus Simian Immunodeficiency virus coated with the envelope proteins, G-protein, from Vesicular Stomatitus virus. This vector is referred to as VSV G-pseudotyped lentivirus, and infects an almost universal set of cells. This tropism is characteristic of the VSV G-protein with which this vector is coated. Many attempts have been made to limit the tropism of viral vectors to one or a few host cell populations. This advance would allow for the systemic administration of a relatively small amount of vector. The potential for off-target cell modification would be limited, as well as many concerns from the medical community. Most attempts to limit tropism have used chimeric envelope proteins bearing antibody fragments. These vectors show great promise for the development of "magic bullet" gene therapies