The Role of Genetics in the Detection of Neurological Diseases: PCR and Cell Culture


ASLAN KOŞAR P.

2nd International Brain Research School, Isparta, Türkiye, 6 - 12 Kasım 2017, cilt.9, sa.3, ss.638-639

  • Yayın Türü: Bildiri / Tam Metin Bildiri
  • Cilt numarası: 9
  • Basıldığı Şehir: Isparta
  • Basıldığı Ülke: Türkiye
  • Sayfa Sayıları: ss.638-639

Özet

The Role of Genetics in the Detection of Neurological Diseases: PCR and Cell Culture Pınar ASLAN KOŞAR Depatment of Medical Biology, Faculty of Medicine, Suleyman Demirel University, Isparta, Turkey The neurogenetics is research of genes that cause neurological disorders and their molecular mechanisms. A clear genetic basis of the most common neuro genetic disorders including some forms of epilepsy, movement disorders, mental retardation, muscular dystrophies and peripheral neuropathies has been established. With the development of new technologies in the field of molecular genetics, our approach to diagnosis and treatment methods of diseases have improved considerably. It is essential that these techniques can be used in the diagnosis and treatment of neurogenetic diseases. These advances in molecular biology have identified more than 200 genes that have contributed to neurological diseases to date. One of the methods used to diagnose neurogenetic diseases is the polymerase chain reaction (PCR). PCR is a common laboratory technique used to make many copies of a particular region of DNA. It is very precise and can be used to amplify, or copy, a specific DNA target from a mixture of DNA molecules. PCR is based on using the ability of DNA polymerase to synthesize new strand of DNA complementary to the offered template strand. Because DNA polymerase can add a nucleotide only onto a preexisting 3'-OH group, it needs a primer to which it can add the first nucleotide. Then, to perform PCR, the DNA template that contains the target is added to a tube that contains primers, free nucleotides, DNA polymerase, and the mixture is placed in a PCR machine. This requirement makes it possible to delineate a specific region of template sequence that the researcher wants to amplify. At the end of the PCR reaction, the specific sequence will be amplificated in billions of copies of times. The resulting PCR products are visualized by agarose gel electrophoresis. Agarose gel electrophoresis is the most effective way of separating DNA fragments of varying sizes ranging from 100 bp to 25 kb and for visualization and purification. To separate DNA using agarose gel electrophoresis. DNA is loaded into pre-cast wells in the gel and a current applied. The phosphate backbone of the DNA (and RNA) molecule is negatively charged, therefore when placed in an electric field, DNA fragments will migrate to the positively charged anode. the negatively charged DNA through an agarose gel matrix toward a positive electrode. Shorter DNA fragments migrate through the gel more quickly than longer ones. Thus, the approximate length of a DNA fragment can be determined by running it on an agarose gel. A suitable voltage for separation of the DNA fragments and ethidium bromide are used to visualize the separated DNA fragments Another method that is used to understand the molecular mechanisms of neurogenetic diseases is cell culture. Cell culture refers to the removal of cells from an animal or plant and their subsequent growth in a favorable artificial environment. The cells may be removed from the tissue directly and disaggregated by enzymatic or mechanical means before cultivation, or they may be derived from a cell line or cell strain that has already been established. Cell culture is one of the major tools used in cellular and molecular biology, providing excellent model systems for studying the normal physiology and biochemistry of cells (e.g., metabolic studies, aging), the effects of drugs and toxic compounds on the cells, and mutagenesis and carcinogenesis. The major advantage of using cell culture for any of these applications is the consistency and reproducibility of results that can be obtained from using a batch of clonal cells.References 

Nörogenetik Hastalıklara Tanısal Yaklaşım ve Kullanılan Yöntemler (2011). Dr. Nerses BEBEK, Tur ki ye Kli nik le ri J NeurolSpecial Topics 4(2):1-6 

Joshi M, Deshpande J. (2010). Polymerase Chain Reaction: Methods, principles and applicatıon. J. IJBR 5:81-97.  

Lee PY, Costumbrado J, Hsu CY, Kim YH. (2012). Agarose gel electrophoresis for the separation of DNA fragments. J Vis Exp. 20: pii: 3923.

The Role of Genetics in the Detection of Neurological Diseases: PCR and Cell Culture Pınar ASLAN KOŞAR Depatment of Medical Biology, Faculty of Medicine, Suleyman Demirel University, Isparta, Turkey The neurogenetics is research of genes that cause neurological disorders and their molecular mechanisms. A clear genetic basis of the most common neuro genetic disorders including some forms of epilepsy, movement disorders, mental retardation, muscular dystrophies and peripheral neuropathies has been established. With the development of new technologies in the field of molecular genetics, our approach to diagnosis and treatment methods of diseases have improved considerably. It is essential that these techniques can be used in the diagnosis and treatment of neurogenetic diseases. These advances in molecular biology have identified more than 200 genes that have contributed to neurological diseases to date. One of the methods used to diagnose neurogenetic diseases is the polymerase chain reaction (PCR). PCR is a common laboratory technique used to make many copies of a particular region of DNA. It is very precise and can be used to amplify, or copy, a specific DNA target from a mixture of DNA molecules. PCR is based on using the ability of DNA polymerase to synthesize new strand of DNA complementary to the offered template strand. Because DNA polymerase can add a nucleotide only onto a preexisting 3'-OH group, it needs a primer to which it can add the first nucleotide. Then, to perform PCR, the DNA template that contains the target is added to a tube that contains primers, free nucleotides, DNA polymerase, and the mixture is placed in a PCR machine. This requirement makes it possible to delineate a specific region of template sequence that the researcher wants to amplify. At the end of the PCR reaction, the specific sequence will be amplificated in billions of copies of times. The resulting PCR products are visualized by agarose gel electrophoresis. Agarose gel electrophoresis is the most effective way of separating DNA fragments of varying sizes ranging from 100 bp to 25 kb and for visualization and purification. To separate DNA using agarose gel electrophoresis. DNA is loaded into pre-cast wells in the gel and a current applied. The phosphate backbone of the DNA (and RNA) molecule is negatively charged, therefore when placed in an electric field, DNA fragments will migrate to the positively charged anode. the negatively charged DNA through an agarose gel matrix toward a positive electrode. Shorter DNA fragments migrate through the gel more quickly than longer ones. Thus, the approximate length of a DNA fragment can be determined by running it on an agarose gel. A suitable voltage for separation of the DNA fragments and ethidium bromide are used to visualize the separated DNA fragments Another method that is used to understand the molecular mechanisms of neurogenetic diseases is cell culture. Cell culture refers to the removal of cells from an animal or plant and their subsequent growth in a favorable artificial environment. The cells may be removed from the tissue directly and disaggregated by enzymatic or mechanical means before cultivation, or they may be derived from a cell line or cell strain that has already been established. Cell culture is one of the major tools used in cellular and molecular biology, providing excellent model systems for studying the normal physiology and biochemistry of cells (e.g., metabolic studies, aging), the effects of drugs and toxic compounds on the cells, and mutagenesis and carcinogenesis. The major advantage of using cell culture for any of these applications is the consistency and reproducibility of results that can be obtained from using a batch of clonal cells.