DNA Extraction

DNA (Deoxyribonucleic Acid) is the complex molecule that contains all the genetic information’s necessary to build and maintain a living being. All living things contain DNA inside their cells. This piece of information is passed on to all generations on reproduction itself. But, in a view of a forensic scientist. DNA is not only a molecule it’s biological evidence from which they may find the perpetrators or someone leads to solving the mystery of the case. From a scene of crime, DNA can be found in many forms such as blood stains, semen, saliva etc. These DNA can’t be seen using our naked human eyes. In order to obtain DNA we need to extract it from the biological sources we found in the scene of crime. This process of extraction is carried out using different methods such as Gel Electrophoresis, Spectrophotometry etc.

The ability to extract DNA from biological stains is a key element in forensic genetics. Attempts to remove blood stains using different type of cleansing methods represent routinely faced forensic problem. DNA could be recovered from cloth after exposure to different types of cleaning products.

Blood, as a physical clue found in many violence cases, has the capacity to provide valuable intelligent evidence for forensic purposes and crime reconstruction. The retrieval of DNA from blood stained cloth provides evidence that can link a criminal or the victim with the scene of crime. DNA techniques are the most advanced and reliable tool for human identification. During the past years, a great number of methods for DNA extraction and typing had been introduced into forensic science, with considerable success. In some cases the perpetrators try to wash off the stain by using different cleaning methods.

Blood is a complex mixture or biological fluid that contains many proteins, enzymes and inorganic compounds. Blood is the predominantly found biological evidence in most of the cases. It can be found in various states or conditions like dried, fresh or even in frozen conditions. In many instances, several types of evidences are recovered in distinct forms or contaminated with any other types of valuable evidences. For forensic investigation purposes, all these evidences have their potential values and often provide links between the victim and accused/suspect.

In case of murder, sexual assault or hit and run cases, a few blood drops are frequently recovered from the scene of occurrence. The forensic biologist helps the investigation agencies in the collection of the evidence from the spot of incidence and analysis of the same in the laboratory. Police submit the evidential samples in the Forensic laboratory with a query of DNA analysis, for cases like murder, attempt to murder, rape, concealment of identity of individual/fragmented body parts or where body is transferred after murder from one place to another. In outdoor crimes such as rape/ sexual assault, biological materials transfer from victim to accused/ culprit or vice versa and to the surroundings.

The place of offence must be fixed by the investigation agencies. Therefore, traces of blood droplets found at crime scene should be processed for DNA profiling. But due to the traced quantity, such samples get consumed in presumptive blood tests, such as Benzidine test and usually not considered for DNA analysis.

In a few cases where the blood is washed off from the crime scene/surface, it becomes one of the most crucial problems faced in forensic examination of criminal cases to detect the blood spot. The surfaces are examined for the presence of blood (a necessity) by use of benzidine or phenolphthalein test. The crucial facet is faced when the blood samples are transferred on clothes in very less quantity. Then, it can be directly tested with benzidine that is competent enough to generate complete DNA profiles.

BENZIDINE is a greyish-yellow to greyish -red, crystalline solid. It is toxic by ingestion, inhalation, and skin absorption. Combustion produces toxic oxides of nitrogen. Benzidine molecular weight 184.242g/mol, an aromatic diamine widely used in industrial processes, that’s why, it is used to prepare other chemicals at few instances, it is used for biological analysis. It is also a powerful carcinogen in many animal species. Some presumptive tests have been described for blood stains recognition, one of the most commonly used tests is Benzidine and its derivative Tetramethylbenzidine (TMB). It has been observed that the administration of benzidine has been shown to produce tumours in liver, hamster liver, and other tissues of exposed animals. In most cases to ingestion or inhalation of benzidine workers in the industry suffered a tremendously increased risk of bladder cancer.

Many investigators have studied the mutagenicity of benzidine and their substitute in short-term tests such as the Ames test. Blood-stains from the surfaces should be lifted in such a manner that contamination with inhibitors could be avoided. In a few cases where blood stains are found on wall plaster or surface and it is impossible to collect the intact spot from the wall then the blood stain should be collected on sterile saline-damp gauze.

DNA analysis becomes more complicated when examination of such samples is conducted after a few months/years from sample collection due to the pendency of cases.

In the present era of modern and advanced technologies, DNA analysing methodology has improved highly. One of the recent and conventional technologies is real time PCR which is a widely used technique for quantitation.

Real time PCR enabled forensic scientists to analyze the sample including blood, saliva, hair etc., even in small quantities and help in exonerating the suspect with the available evidence. This technique helps in qualitative, quantitative analysis of DNA. It also indicates the presence of inhibitors in the samples. By using this technique of DNA quantitation in forensic samples, forensic scientists can determine how much DNA is present in the sample and how much we must use for conventional PCR. A required number of amplicons is mandatory for complete DNA profiling, otherwise split peaks, false peaks or off ladder alleles will be in the profile. Inhibitors in the sample may directly bind to DNA or interact with DNA polymerases, Sometimes stains cannot be seen in that cases we use UV detectors or UV lights to locate any stains present and we cannot confirm the stain is blood so we conduct some presumptive tests followed by confirmatory tests like LMG Test, Benzidine Test, Kastle-Meyer test (presumptive tests), Takayama Test, Teichmann Test (confirmatory test) etc.

(This Article is a part of Dissertation work on “Influence of tampered cloth evidence by detergent” by Anita Thomas.)

About The Author

Anita Thomas is currently pursuing a bachelor’s degree in The Degree Of B.Voc Applied Microbiology And Forensic Science.

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