Biotechnology Tools in Forensic Science

Biotechnology is one of the branch that is contributing in forensic science. There are many tools in biotechnology that helps in solving cases in forensic.

1. Biotechnology tools in forensic science Malsha Kularathna BSc(Hons)Biotechnology (UG) AFRS - Volunteer

2. Content Introduction DNA Sequencing PCR Machine Gel Electrophoresis Apparatus UV – VIS Spectroscopy Microarray 2

3. Introduction What is Forensic Biotechnology? Forensic biotechnology refers to the application of biotechnology techniques and methods for investigating legal cases. This field combines principles of biology, genetics, and molecular biology. Forensic biotechnology can be used in a variety of contexts, including criminal investigations, paternity testing, and identification of human remains. Some common techniques and tools used in forensic biotechnology include, DNA Sequencing machine PCR (polymerase chain reaction) machine Gel Electrophoresis Apparatus UV-VIS Spectrometer Microarray 3

4. 01. DNA Sequencing DNA sequencing is the process of determining the exact order of nucleotides in a DNA sample. In forensic science DNA sequencing is used to determine paternity, and analyse genetic mutations. The steps involved in DNA sequencing using a DNA sequencing machine are as follows: Sample Preparation Library Preparation Loading the DNA Sequencer Sequencing Data Analysis Sample Preparation: The DNA sample is extracted from the biological material, purified, and quantified to ensure that there is enough DNA for sequencing. The sample is then fragmented into smaller pieces for sequencing. Library Preparation: The fragmented DNA is ligated with adapters that allow it to be amplified and sequenced. The resulting library is amplified through PCR to create multiple copies of the DNA fragments . 4

5. Loading the DNA Sequencer: The DNA library is loaded into the DNA sequencer, where it is immobilized on a flow cell or a sequencing chip. Sequencing: The sequencing machine adds nucleotides to the immobilized DNA fragments, one at a time, and records the fluorescent signals generated as each nucleotide is incorporated. The sequencing machine then identifies the specific sequence of each fragment based on the order of the fluorescent signals. Data Analysis: The raw sequencing data is then processed using specialized software to remove errors, filter out low-quality reads, and assemble the reads into a complete sequence. Figure 01 – DNA Sequencing Machine 5

6. 02. PCR Machine Polymerase chain reaction (PCR) machines are used to amplify small amounts of DNA into larger quantities that can be analysed. PCR is often used to generate DNA profiles from very small or degraded samples, such as those found in bones or hair. Figure 02 – PCR Machine Figure 03 – Parts of PCR Machine 6

7. The PCR method requires the following four reagents or chemicals: DNA or RNA sample DNA primers:- short single-stranded DNA molecule that promotes synthesis of a complementary strand of nucleotides DNA polymerase:- an enzyme that aids in the synthesis of a complementary strand of DNA Nucleotide solution mix containing The PCR process has 4 steps: Denaturation Annealing Extension Repeated cycles Denaturation: The double-stranded DNA is heated to a high temperature (typically 94-98°C) to separate the two strands of DNA. Annealing: The temperature is lowered to allow the primers to anneal (bind) to the complementary single-stranded DNA templates. The primers are short pieces of DNA that are designed to bind to the specific sequence of DNA that is to be amplified. 7

8. Extension: The temperature is raised to a temperature that is optimal for the DNA polymerase enzyme to add nucleotides to the 3' end of each primer, extending the DNA strands. The DNA polymerase enzyme extends the primers in a 5' to 3' direction, using the single-stranded DNA template as a template for the new DNA strand. Repeated cycles: The denaturation, annealing, and extension steps are repeated multiple times (usually 20-40 cycles) to amplify the DNA segment of interest exponentially. The number of DNA segments is doubled with each cycle of PCR, resulting in a very large number of copies of the targeted DNA segment. 8

9. In forensic science, PCR is used for various applications . DNA profiling : This technique is used in forensic investigations to identify suspects or to match DNA samples collected from a crime scene. Forensic paternity testing : This technique is used in cases where the identity of the biological father needs to be established. Detection of pathogens : PCR is used to detect the presence of pathogens such as viruses or bacteria in forensic samples. Figure 04 – DNA paternity testing 9

10. 03. Gel Electrophoresis Apparatus Gel electrophoresis is a laboratory technique used to separate & analyse mixtures of biological macromolecules such as DNA, RNA & proteins based on their size, charge & shape. Figure 05 – Parts of Gel Electrophoresis 10

11. Procedure of Gel electrophoresis apparatus Preparation of the gel Loading the gel into the electrophoresis tank Loading the biological sample Applying the electric field Running the electrophoresis Staining the gel Documenting the results Figure 06 – Procedure of Gel Electrophoresis 11

12. Gel electrophoresis is used in forensic science to analyse DNA evidence , such as in criminal investigations, analysis of mitochondrial DNA and paternity testing . 12

13. 04. UV-VIS Spectroscopy The best analytical method for analysing a variety of evidence, including bodily fluids, narcotics, and trace components discovered at the scene of the crime, is spectrophotometry. Identification of smokeless powder using spectroscopy When no weapon is discovered in a shooting crime, smokeless powder will be the key piece of forensic evidence. Figure 06 - Composition of smokeless powder 13

14. A UV-Vis spectrophotometer for fiber analysis Fiber is a type of evidence found at crime scenes, similar to hair. Fibers as evidence have probative value in forensic science because they establish linkages or associations between the suspect and the victim. Figure 07 – Fiber as evidence 14

15. Spectroscopy technique to analyze automotive paints Paint fragments found at the scene of the crime can be used as forensic evidence in cases of automobile accidents, vandalism, and burglaries. This evidence aids in identifying crimes. Spectroscopy for forensic toxiology The investigation of biological materials to identify toxins or medications is known as forensic toxicology. Toxicology provides the information what type of substance and the amount of substance used which is potential effect for the person illness or death Figure 08- UV Spectroscopy 15

16. 05.Microarrays A microarray is a laboratory tool used to simultaneously monitor the expression of thousands of genes. They can be used in forensic science to identify genetic variations and compare DNA samples. Figure 09 - Microarray 16

17. Microarrays can be used for a wide range of forensic applications, including: DNA identification: Microarrays can be used to identify individuals by comparing their DNA profiles to those in a database. Ancestry testing: Microarrays can be used to identify an individual's ancestry by comparing their DNA to reference populations. Paternity testing: Microarrays can be used to determine paternity by comparing the DNA of an alleged father to that of a child. Forensic DNA phenotyping: Microarrays can be used to predict physical characteristics of an individual based on their DNA. Figure 10 – Microarray technique 17

18. References DNA Sequencing. (n.d.). Genome.gov. https://www.genome.gov/genetics-glossary/DNA-Sequencing Wikipedia contributors. (2023). DNA sequencing. Wikipedia. https://en.wikipedia.org/wiki/DNA_sequencing Karki, P. (2022). PCR Machine- Principle, Parts, Steps, Types, Uses, Examples. Microbe Notes. https://microbenotes.com/pcr-machine-principle-parts-steps-types-uses-examples/ gel electrophoresis  | Learn Science at Scitable . (n.d.). https://www.nature.com/scitable/definition/gel-electrophoresis-286/ SMACgig World. (2022). Spectroscopy and Forensics. www.smacgigworld.com. https://www.smacgigworld.com/blog/spectroscopy-and-forensics.php microarray  | Learn Science at Scitable . (n.d.). https://www.nature.com/scitable/definition/microarray-202/ Nhgri. (2019). DNA Microarray Technology Fact Sheet. Genome.gov. https://www.genome.gov/about-genomics/fact-sheets/DNA-Microarray-Technology 18

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