Adrienne Dellwo, Fibromyalgia & CFS Blog
ABSTRACT
NEWSBRIEF: A virus suspected of triggering some cases of chronic fatigue syndrome (CFS or ME/CFS) can be passed genetically from parent to child, according to an article published in the journal Pediatrics.
Experts used to believe that mothers passed human herpesvirus 6 (HHV-6) to their babies through blood exchanged during childbirth. HHV-6 causes roseola in infected children. New evidence shows the virus can come from either parent in the genetic material.
Nearly everyone is infected with HHV-6 at some point in their lives, but researchers are now investigating what it means to have a virus integrated into your genetic matter -- whether it can activate and cause problems, or cause an immune response just by being there.
Guide note: Could this be causing a genetic predisposition to chronic fatigue syndrome or autoimmune diseases? If so, genetic testing may someday be able to diagnose the condition and reveal who is at risk for it.
Nov 28, 2008
HHV-6 is Passed to Children Through DNA
Brief to Genetic Mapping
What is genetic mapping?
Developing new and better tools to make gene hunts faster, cheaper and practical for any scientist was a primary goal of the Human Genome Project (HGP).
One of these tools is genetic mapping, the first step in isolating a gene. Genetic mapping - also called linkage mapping - can offer firm evidence that a disease transmitted from parent to child is linked to one or more genes. It also provides clues about which chromosome contains the gene and precisely where it lies on that chromosome.
Genetic maps have been used successfully to find the single gene responsible for relatively rare inherited disorders, like cystic fibrosis and muscular dystrophy. Maps have also become useful in guiding scientists to the many genes that are believed to interact to bring about more common disorders, such as asthma, heart disease, diabetes, cancer and psychiatric conditions.
How do researchers create a genetic map?
To produce a genetic map, researchers collect blood or tissue samples from family members where a certain disease or trait is prevalent. Using various laboratory techniques, the scientists isolate DNA from these samples and examine it for the unique patterns of bases seen only in family members who have the disease or trait. These characteristic molecular patterns are referred to as polymorphisms, or markers.
Before researchers identify the gene responsible for the disease or trait, DNA markers can tell them roughly where the gene is on the chromosome. This is possible because of a genetic process known as recombination. As eggs or sperm develop within a person's body, the 23 pairs of chromosomes within those cells exchange - or recombine - genetic material. If a particular gene is close to a DNA marker, the gene and marker will likely stay together during the recombination process, and be passed on together from parent to child. So, if each family member with a particular disease or trait also inherits a particular DNA marker, chances are high that the gene responsible for the disease lies near that marker.
The more DNA markers there are on a genetic map, the more likely it is that one will be closely linked to a disease gene - and the easier it will be for researchers to zero-in on that gene. One of the first major achievements of the HGP was to develop dense maps of markers spaced evenly across the entire collection of human DNA.
Markers themselves usually consist of DNA that does not contain a gene, however they can tell a researcher the identity of the person a DNA sample came from. This makes markers extremely valuable for tracking inheritance of traits through generations of a family, and markers have also proven useful in criminal investigations and other forensic applications.
Although there are several different types of genetic markers, the type most used on genetic maps today is known as a microsatellite map. However, maps of even higher resolution are being constructed using single-nucleotide polymorphisms, or SNPs (pronounced "snips"). Both types of markers are easy to use with automated laboratory equipment, so researchers can rapidly map a disease or trait in a large number of family members.
The development of high-resolution, easy-to-use genetic maps, coupled with the HGP's successful sequencing and physical mapping of the entire human genome, has revolutionized genetics research. The improved quality of genetic data has reduced the time required to identify a gene from a period of years to, in many cases, a matter of months or even weeks. Genetic mapping data generated by the HGP's laboratories is freely accessible to scientists through databases maintained by the National Institutes of Health and the National Library of Medicine's National Center for Biotechnology Information.
Nov 23, 2008
DNA Microchip Technology
What is a DNA microchip?
Scientists know that a mutation - or alteration - in a particular gene's DNA often results in a certain disease. However, it can be very difficult to develop a test to detect these mutations, because most large genes have many regions where mutations can occur. For example, researchers believe that mutations in the genes BRCA1 and BRCA2 cause as many as 60 percent of all cases of hereditary breast and ovarian cancers. But there is not one specific mutation responsible for all of these cases. Researchers have already discovered over 800 different mutations in BRCA1 alone.
The DNA microchip is a mew tool used to identify mutations in genes like BRCA1 and BRCA2. The chip, which consists of a small glass plate encased in plastic, is manufactured somewhat like a computer microchip. On the surface, each chip contains thousands of short, synthetic, single-stranded DNA sequences, which together add up to the normal gene in question.
What is a DNA microchip used for?
Because chip technology is still relatively new, it is currently only a research tool. Scientists use it to conduct large-scale population studies - for example, to determine how often individuals with a particular mutation actually develop breast cancer.
As we gain more insight into the mutations that underlie various diseases, researchers will likely produce new chips to help assess individual risks for developing different cancers as well as heart disease, diabetes and other diseases.
How does a DNA microchip work?
To determine whether an individual possesses a mutation for BRCA1 or BRCA2, a scientist first obtains a sample of DNA from the patient's blood as well as a control sample - one that does not contain a mutation in either gene.
The researcher then denatures the DNA in the samples - a process that separates the two complementary strands of DNA into single-stranded molecules. The next step is to cut the long strands of DNA into smaller, more manageable fragments and then to label each fragment by attaching a fluorescent dye. The individual's DNA is labeled with green dye and the control - or normal - DNA is labeled with red dye. Both sets of labeled DNA are then inserted into the chip and allowed to hybridize - or bind - to the synthetic BRCA1 or BRCA2 DNA on the chip. If the individual does not have a mutation for the gene, both the red and green samples will bind to the sequences on the chip.
If the individual does possess a mutation, the individual's DNA will not bind properly in the region where the mutation is located. The scientist can then examine this area more closely to confirm that a mutation is present.
Last Reviewed: November 3, 2008 （Source：National Human Genomic Program）
DNA Microchips
Perhaps the best-known type of surface attachment involves DNA oligonucleotide microarrays (so called gene chips). Their importance to the study of genetics was illustrated by the appearance in (1999) of an entire supplemental issue of Nature Genetics dedicated to the subject. These chips have been used to enable rapid sequencing of DNA, for the detection of RFLPs (Hacia & Collins, 1999) , for monitoring the presence of microorganisms in the food industry (Kuipers, 1999; Kuipers et al., 1999) , or the environment (Guschin et al., 1997; Oh & Liao, 2000) and to assay gene expression in a wide range of organisms (Harrington et al., 2000) . More recently, chips have been manufactured with a three-dimensional array based on the use of a polyacrylamide matrix (Guschin et al., 1997) . These are likely to lead to faster screening techniques and the use of larger “databases” of genes on the chips for expression studies. Microchip development has also spawned a vast collection of associated industries producing clone-sets, arrayers, scanners and software for data analysis (Epstein & Butlow, 2000) . It seems likely that single chips will get smaller, moving toward the nanoscale and the detection/use of single DNA molecules.
Currently, there are two major variants of DNA chip technology based on the type of immobilised DNA. In the first format, the immobilised probe consists of cDNA (500-5000bp DNA fragments produced by copying expressed genes) spotted onto a surface using robotics. The second format, consists of 20-25-mer oligonucleotides (or peptide nucleic acid - PNA) synthesised in situ (such as the gene chips produced by Affymetrix), or by conventional synthesis and then immobilised. The oligonucleotides are of known DNA sequence allowing detection of complementary sequences. Therefore, the DNA on the chip acts as a probe in a hybridisation reaction (Souteyrand et al., 2000; Southern et al., 1999) . The arrays come in various sizes from macroarrays to microarrays and beyond. Macroarrays are generally produced by hand, or using simple robots to pipette DNA solutions onto prepared surfaces and contains “tens to hundreds” of spots over an area of 500 mm or more. Microarrays contain many thousands of spots over an area of less than 500 mm.
Gene chips for cancer detection
DNA microchips have been used extensively for research purposes, but they also have a powerful use as a biosensor. For example, the onset of prostate cancer involves the differential expression of approximately 500 genes. A prototype chip for prostate cancer detection is already under development by the biotechnology firm Darwin Molecular (Bothell, Washington USA). They predict that within a year they will have a system that can distinguish slow-growing cancers that do not require immediate treatment and those aggressive tumours that require surgery (Zajtchuk, 1999).
Detection of hybridisation usually involves the use of PCR amplified, fluorescently-tagged DNA. Generally, for gene expression studies, this involves amplification of a pair of mRNA samples (i.e. a control and an experimental sample) using different fluorophores for each sample. Identical amounts of the amplified cDNA are hybridised to the same DNA chip and fluorescent intensities compared (Winzeler et al., 1999) . The use of two fluorophores overcomes problems associated with different amounts of probe DNA deposited at the specific spots on the chip.
The first consideration in the design of a DNA chip is the support material and the production of simple arrays in a research environment was discussed by Cheung et al. (1999) . Most usually, this involves the use of glass. However, recently silicon has also been employed as a support allowing the use of its semiconducting properties (Souteyrand et al., 2000) . Electrochemical deposition of oligonucleotides, via pyrrole groups is used to attach the DNA to the surface of electrodes (Livache et al., 1998a) . These electrodes can interact with the silicon allowing for the development of new techniques for detecting hybridisation based on the semiconductor properties of the silicon. A typical 128-sample grid is shown in Figure 17. The chip uses polarisation of the electrodes (Souteyrand et al., 2000) to prevent diffusion, allowing the samples to concentrate ensuring hybridisation occurs in a few seconds. After hybridisation, a short burst of the opposite polarisation will release weakly bound molecules improving signal-noise ratios. Finally, the charge on the DNA strands allows detection of hybridisation without using fluorophores, through the field-effects on the electrodes.
Perhaps the company most associated with DNA microchips, Affymetrix, believes that the capacity of DNA microchips is following Moore’s Law, which predicts a two-fold increase in computer chip memory every 18 months. Affymetrix’s prototype chips held 20,000 oligonucleotide DNA probes. By 1997, this number had increased to 65,000 probes and it has recently introduced a chip with 400,000 probes. Ten such chips could scan the entire human genome for ‘malfunctioning’ genes.
（Source：Keith Firman, Nanonet)
Deoxyribonucleic Acid (DNA)
What is DNA?
We all know that elephants only give birth to little elephants, giraffes to giraffes, dogs to dogs and so on for every type of living creature. But why is this so?
The answer lies in a molecule called deoxyribonucleic acid (DNA), which contains the biological instructions that make each species unique. DNA, along with the instructions it contains, is passed from adult organisms to their offspring during reproduction.
DNA is found inside a special area of the cell called the nucleus. Because the cell is very small, and because organisms have many DNA molecules per cell, each DNA molecule must be tightly packaged. This packaged form of the DNA is called a chromosome.
DNA spends a lot of time in its chromosome form. But during cell division, DNA unwinds so it can be copied and the copies transferred to new cells. DNA also unwinds so that its instructions can be used to make proteins and for other biological processes.
Researchers refer to DNA found in the cell's nucleus as nuclear DNA. An organism's complete set of nuclear DNA is called its genome.
Besides the DNA located in the nucleus, humans and other complex organisms also have a small amount of DNA in cell structures known as mitochondria. Mitochondria generate the energy the cell needs to function properly.
In sexual reproduction, organisms inherit half of their nuclear DNA from the male parent and half from the female parent. However, organisms inherit all of their mitochondrial DNA from the female parent. This occurs because only egg cells, and not sperm cells, keep their mitochondria during fertilization.
DNA is made of chemical building blocks called nucleotides. These building blocks are made of three parts: a phosphate group, a sugar group and one of four types of nitrogen bases. To form a strand of DNA, nucleotides are linked into chains, with the phosphate and sugar groups alternating.
The four types of nitrogen bases found in nucleotides are: adenine (A), , thymine (T), guanine (G) and cytosine (C). The order, or sequence, of these bases determines what biological instructions are contained in a strand of DNA. For example, the sequence ATCGTT might instruct for blue eyes, while ATCGCT might instruct for brown.
Each DNA sequence that contains instructions to make a protein is known as a gene. The size of a gene may vary greatly, ranging from about 1,000 bases to 1 million bases in humans.
The complete DNA instruction book, or genome, for a human contains about 3 billion bases and about 20,000 genes on 23 pairs of chromosomes.
DNA contains the instructions needed for an organism to develop, survive and reproduce. To carry out these functions, DNA sequences must be converted into messages that can be used to produce proteins, which are the complex molecules that do most of the work in our bodies.
How are DNA sequences used to make proteins?
DNA's instructions are used to make proteins in a two-step process. First, enzymes read the information in a DNA molecule and transcribe it into an intermediary molecule called messenger ribonucleic acid, or mRNA.
Next, the information contained in the mRNA molecule is translated into the "language" of amino acids, which are the building blocks of proteins. This language tells the cell's protein-making machinery the precise order in which to link the amino acids to produce a specific protein. This is a major task because there are 20 types of amino acids, which can be placed in many different orders to form a wide variety of proteins.
The German biochemist Frederich Miescher first observed DNA in the late 1800s. But nearly a century passed from that discovery until researchers unraveled the structure of the DNA molecule and realized its central importance to biology.
For many years, scientists debated which molecule carried life's biological instructions. Most thought that DNA was too simple a molecule to play such a critical role. Instead, they argued that proteins were more likely to carry out this vital function because of their greater complexity and wider variety of forms.
The importance of DNA became clear in 1953 thanks to the work of James Watson, Francis Crick, Maurice Wilkins and Rosalind Franklin. By studying X-ray diffraction patterns and building models, the scientists figured out the double helix structure of DNA - a structure that enables it to carry biological information from one generation to the next.
Scientist use the term "double helix" to describe DNA's winding, two-stranded chemical structure. This shape - which looks much like a twisted ladder - gives DNA the power to pass along biological instructions with great precision.
To understand DNA's double helix from a chemical standpoint, picture the sides of the ladder as strands of alternating sugar and phosphate groups - strands that run in opposite directions. Each "rung" of the ladder is made up of two nitrogen bases, paired together by hydrogen bonds. Because of the highly specific nature of this type of chemical pairing, base A always pairs with base T, and likewise C with G. So, if you know the sequence of the bases on one strand of a DNA double helix, it is a simple matter to figure out the sequence of bases on the other strand.
DNA's unique structure enables the molecule to copy itself during cell division. When a cell prepares to divide, the DNA helix splits down the middle and becomes two single strands. These single strands serve as templates for building two new, double-stranded DNA molecules - each a replica of the original DNA molecule. In this process, an A base is added wherever there is a T, a C where there is a G, and so on until all of the bases once again have partners.
In addition, when proteins are being made, the double helix unwinds to allow a single strand of DNA to serve as a template. This template strand is then transcribed into mRNA, which is a molecule that conveys vital instructions to the cell's protein-making machinery.
Last Reviewed: October 1, 2008
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