DNA profiling How is it carried out?

The process begins with a sample of an individual's DNA (typically called a "reference sample"). The most desirable method of collecting a reference sample is the use of a buccal swab, as this reduces the possibility of contamination. When this is not available (e.g. because a court order may be needed and not obtainable) other methods may need to be used to collect a sample of blood, saliva, semen, or other appropriate fluid or tissue from personal items (e.g. toothbrush, razor, etc.) or from stored samples (e.g. banked sperm or biopsy tissue). Samples obtained from blood relatives (biological relative) can provide an indication of an individual's profile, as could human remains which had been previously profiled.

A reference sample is then analyzed to create the individual's DNA profile using one of a number of techniques, discussed below. The DNA profile is then compared against another sample to determine whether there is a genetic match.

There are many methods that can be used to carry this out. One of which is restriction fragment length polymorphism (RFLP) analysis. It involve restriction enzyme digestion, followed by Southern blot analysis. Although polymorphisms can exist in the restriction enzyme cleavage sites, more commonly the enzymes and DNA probes were used to analyze VNTR loci. However, the Southern blot technique is laborious, and requires large amounts of undegraded sample DNA. Also, Karl Brown's original technique looked at many minisatellite loci at the same time, increasing the observed variability, but making it hard to discern individual alleles (and thereby precluding parental testing).

Variations of VNTR allele lengths in 6 individuals. Another method is the PCR analysis. With the invention of the polymerase chain reaction (PCR) technique, DNA profiling took huge strides forward in both discriminating power and the ability to recover information from very small (or degraded) starting samples. PCR greatly amplifies the amounts of a specific region of DNA, using oligonucleotide primers and a thermostable DNA polymerase.

Below is an article showing how DNA profiling has benefited the society. By Sang Mi Park

Deoxyribonucleic Acid (DNA) is the fundamental building block of all living matter. Functioning as the "blue print" for living structures, DNA is present in all cells and most virii. DNA carries all the information that is needed for protein synthesis and replication of cells. In living organisms, DNA is organized in chromosomes, and it is located in the nucleus of each cell. One of the cases in which DNA helped to solve a crime was the Linda Mann case.

In 1983, Linda Mann, 15, was found murdered. The killer had left a small sample of his semen, which was tested. Four years later, Dawn Ashcroft was found raped and strangled to death. The police believed that the same man had committed both crimes. A dishwasher was a suspect in the crime, and later confessed to it.

Later on in the investigation, the police utilized a new method (DNA fingerprinting). The technique was developed by an English geneticist by the name of Alec Jeffreys. It allowed the investigators to compare the suspect's DNA with the DNA from the semen found at the crime scene. After the results came in, the police were surprised to see that the DNA testing proved that the dishwasher was innocent of both murders.

Later on, a young woman who managed a local bakery said that she had overheard a man telling another man that he had paid someone to go in his place to give blood in his name (Colin Pitchfork). He had a record and had been arrested several times for indecent exposure. The police confronted Colin Pitchfork with the murder accusations, and being convinced that the DNA identification would convict him, Pitchfork admitted to the murders. DNA fingerprinting, a relatively new procedure, had helped to solve a crime.

http://www.bxscience.edu/publications/forensics/articles/dna/h-dna08.htm http://library.thinkquest.org/17049 Coleman, Howard. DNA in the Courtroom: A Trial Watcher's Guide New York: Scholastic, 1999 DNA technology is constantly evolving. New techniques, new loci and the ability to anaylse smaller samples with increased automation promise faster and more discriminating results for the presentaton of forensic evidence incourt.

http://nzic.org.nz/ChemProcesses/biotech/12D.pdf

Genes make up the blueprint for our bodies, governing factors such as growth, development and functioning. Almost every cell in the human body contains a copy of the blueprint, stored inside a special sac called the nucleus. The estimated 30,000 genes are beaded along tightly bundled strands of a chemical substance called deoxyribonucleic acid (DNA). DNA profiling is a way of establishing identity and is used in a variety of ways, such as finding out whether twins are fraternal or identical. DNA samples are usually obtained from blood.

While DNA contains material common to all humans, some portions are unique to each individual. These portions, or regions, contain two genetic types (alleles) that are inherited from the person’s mother and father. A person’s DNA profile is made by investigating a number of these regions. In a paternity test, for example, the mother’s DNA profile is compared with the child’s to find which half was passed on by the mother. The other half of the child’s DNA is then compared with the alleged father’s DNA profile. If they don’t match, the ‘father’ is excluded, which means he isn’t the father of that child. If the DNA profiles match, the ‘father’ is not excluded - which means there is a high probability (more than 99 per cent) that he is the father. DNA tests such as this can’t offer 100 per cent proof.

Some of the advantages of DNA profiling include:

* DNA tests can be applied to any human sample that contains cells with nuclei, such as saliva, semen, urine and hair.

* DNA is hardy, and resists degeneration even after contamination with chemicals or bacteria.

* The ability of DNA profiling to exclude a suspect means the police are able to confidently drop that line of enquiry and continue their investigation down other avenues. Between 1989 and 1996, the FBI used genetic testing in about 10,000 sexual assault cases; in 2,000 of those cases, the prime suspect was discovered to have not committed the crime. Without genetic testing, it can be assumed that some of these men would have been convicted. In fact, many prison inmates have appealed their conviction after spending years in jail, and have been discovered to be innocent.

Contrary to public belief, DNA profiling isn’t infallible. Critics point out various problems and limitations, including:

* New DNA profiling technologies can give incorrect results, due to errors such as cross-contamination of samples.

* Older DNA profiling technologies are more prone to errors, which could give false-negative or false-positive results.

* DNA profiles can only offer statistical probability (for example, one in a million), rather than absolute certainty.

SINGAPORE SCIENTISTS’ FINDING ON EARLY EMBRYONIC DEVELOPMENT HAS DIRECT IMPACT ON REGENERATIVE MEDICINE AND ASSISTED REPRODUCTION

Scientists at the Genome Institute of Singapore (GIS) have recently generated significant single cell expression data crucial for a detailed molecular understanding of mammalian development from fertilization to embryo implantation, a process known as the preimplantation period. The knowledge gained has a direct impact on clinical applications in the areas of regenerative medicine and assisted reproduction.
This study, published in Developmental Cell on April 20, 2010, is the first of its kind to apply single cell gene expression analysis of many genes to hundreds of cells in a developmental system.
Using the new BioMark microfluidic technology and the mouse preimplantation embryo as a model, the scientists were able to study the expression of 48 genes from individual cells and applied this to analyze over 600 individual cells from the 1-cell to the 64-cell stage of preimplantation development. This high throughput single cell research methodology provides the scientists with the ability to detect dynamic patterns in cellular behaviour, which is unprecedented in the field. Significantly, the findings of the study resolves some of the arguments pertaining to cellular differentiation events and places fibroblast growth factor signalling as the primary event in the later cell fate decisions.
Executive Director at the GIS, a biomedical research institute of the Agency for Science, Technology and Research (A*STAR), Professor Edison Liu said,
“This remarkable work by Guoji Guo, Mikael Huss, Paul Robson and colleagues uses new microgenomic technologies to map, over time, how a single cell decides to permanently become different parts of an embryo. Within one division, cells commit to specific developmental lineages by expressing defined sets of genes. This research now opens the possibility of assessing the genetic triggers for fate determination of individual cells in developmental time. On another level, this work highlights the importance of new microtechnologies in advancing the understanding of early embryonic events. “
Professor Davor Solter, Senior Principal Investigator of the Institute of Medical Biology, A*STAR, added, “This is a real technological tour de force. The authors investigated changes in expression of multiple genes on the single cell level during preimplantation mouse development. They clearly demonstrated gradual and stochastic lineage allocation and absence of predetermination. These results conclusively resolved one of the hotly debated issues in mammalian development and provided important new insight into the mechanism which regulates early development in mammals.”
"These are important findings. The team at GIS provided a new look into the complex and little-understood process of early embryo development. It also demonstrates the power of single cell gene expression. It is clear that individual cells and small groups of cells behave differently than the aggregate population, and these differences are key to understanding the biology of the system as a whole." said Gajus Worthington, president and chief executive officer of Fluidigm. "It always provides a special thrill when researchers use the capabilities of Fluidigm's technology to bring insight to the body of scientific knowledge."
The Preimplantation period involves the first cellular differentiation events in mammalian development including the formation of pluripotent cells from where embryonic stem (ES) cells are derived. Being one of the simplest mammalian developmental systems to study, it can provide comprehensive understanding of the complex molecular control of reprogramming and cell fate decisions.

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