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Genetics Research

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The Future of Genetics

The HGP began in 1990, it is a 13-year effort coordinated and funded by the U.S. Department of Energy and the National Institutes of Health. The Human Genome Project's goals are to identify all the 100,000 genes in human DNA; determine the sequences of the 3 billion chemical base pairs that make up human DNA; store this information in databases; develop tools for data analysis; transfer related technologies to the private sector; and address the ethical, legal, and social issues (ELSI) that may arise from the project. A working draft of the human sequence was completed earlier this year, 2000. The U.S. Human Genome Project (HGP), composed of the DOE and NIH Human Genome Programs, is the national coordinated effort to characterize all human genetic material by determining the complete sequence of the DNA in the human genome. The HGP's ultimate goal is to discover all the more than 80,000 human genes and render them accessible for further biological study. To facilitate the future interpretation of human gene function, parallel studies are being carried out on selected model organisms, such as Drosophilia Melanogaster and Caenorhabditis elegans. According to the department of energy program report, a perfect draft of the human sequence is due in 2003.

Some of the ways that geneticists use to map the Human Gene are Atomic Force Microscopy of Biochemically Tagged DNA, Intracellular Flow Karyotyping, and Electrotransformation for Introducing DNA into Industrial Bacilli

Intracellular flow karyotyping appears to be a feasible and beneficial method for analyzing karyotype aberrations from individual cells using flow cytogenetics. The flow karyotyping method allows quantification of chromosomal DNA by flow cytometry and thus analysis of chromosomal aberrations on chromosome suspensions. Amounts of data providing statistical significance can be collected quickly and the approach allows accurate mapping of chromosomal DNA composition. The limitation of the method is at the cellular level of analysis, which is an impossibility to detect low-frequency or heterogeneous events, with this method. The aim of this intracellular flow karyotyping project is improving the technology to extend the method to the analysis of karyotype aberrations from individual cells.

This technology might be especially useful for the detection and quantification of heterogeneous abnormalities. Chromosomal changes of this type occur through ionising radiation exposure and are involved in karyotype instability and tumorigenesis. This approach will be investigated both for biological dosimetry purposes, especially in low-dose contexts (count of abnormal cells, count of abnormalities per cell) and for research purposes (karyotype instability known as tumorigenesis).

Preliminary results demonstrating the feasibility were obtained using hydrodynamic destruction of mitotic cells by capillary flow, high gradient devices and monovariate (DNA quantification) flow karyotyping. This approach of cell membrane destruction will be optimised and alternative methods (particularly ultrasonic disintegration) developed. The intracellular staining method of chromosomes with DNA specific fluorochromes will be improved especially for dual parameter (DNA content and base pair composition quantification) intracellular flow karyotype analysis. The method will be adapted for modern serial flow, cytometer systems (first step: partners' equipment). The development of new algorithms and computer programs for data interpretation is in progress. In parallel to the technical improvements pilot research using different human cell line models will be conducted to investigate the method's parameters.

Another way used to map genes is Atomic Force Microscopy of Biochemically Tagged DNA. According to the 1998 cytometry report by V Zenin, this process uses small DNA fragments of a known length. They are made using a polymerase chain reaction. These frag-ments contain biotin molecules, usually vitamin H, covalently attached to each end. Then the DNA is labeled with streptavidin. This tetrameric complex was expected to bind up to four DNA molecules via their attached biotin molecules. The DNA is then imaged with atomic force microscopy (AFM). Images revealed the protein at the end of the DNA strands as well as the presence of dimers, trimers, and tetramers of DNA bound to a single protein. Imaging time was about 1 min.

The DOE Program report states "With these results, we believe we have shown that AFM does have sufficient resolution to map DNA. In its simplest form, mapping involves measuring the physical distance between two points of DNA. In this experiment we have demonstrated the ability of AFM to perform this task by attaching a large protein marker to genetically engineered pieces of human DNA and using AFM to locate the marker and measure the known length from the protein to the other end of the DNA."

Electrotransformation for Introducing DNA into Industrial Bacilli is perhaps one of the most interesting techniques developed by the Human Genome Project, because of its industrial uses. According to PATN, ""Electrotransformation" (ET) is the process whereby genetic material is taken up into a host cell and subsequently becomes part of the host cell genome by utilization of long-term, non-pulsed electric current for a sufficient time to effect transformation." The project is based on experiments revealing the unexpected roles of electric field intensity for cell-wall permeability and the dependence of pulse shape on ET efficiency. The observations became apparent through use of a specially constructed apparatus in which electric pulse parameters were independent of cell suspension; thus allowing pulse shapes to vary. Geneticists first use enzymes to "cut out" small sections of DNA. Then they insert the DNA in the place of the DNA of the bacilli. The bacilli then produce what the small section of DNA tells it to produce. Thus the advent of industrially interesting microbes.

According to IFS News, genetically produced "human" insulin was introduced in 1982. Prior to this highly purified bovine or porcine insulin had been used which had successfully removed the skin and injection site problems, which was common with the older, less pure insulin. In this case, scientists used electrotransformation. When they had accomplished this feat, the world's insulin supply became much higher as well as less costly, and the promise of dollars drew new companies.

Celera Genomics Corp. is the Human Genome Project's larges competitor in the United States. Celera



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