Chemicals used in pcr

Polymerase chain reaction, PCR, chemicals used in PCR, Ingredients used in PCR, Deionized water, PCR Buffer, MgCl2, dNTPS, types of dNTPS, Taq DNA polymerase, Primers, Amplicon, Molecular method.

1. CHEMICALS USED IN PCR BY HASNAT TARIQ

2. CONTENTS DEIONIZED WATER PCR BUFFER MgCL2 dNTPs PRIMERS TAQ POLYMERASE TARGET DNA SEQUENCE 2

3. DEIONIZED WATER Also called double distilled water . 11.4 μl is used per sample in PCR Provides physiochemical environment . Also d ilute chemical reagents. 3

4. 2. PCR BUFFER 10 X buffer is used. 2μl is used per sample. Maintains pH . Provides physiochemical environment . Optimize conditions like salt concentration and pH to enable Taq polymerase to work efficiently. 2 types : KCL Facilitate template primer binding. 2. (NH4)2SO4 Increase primer specificity. 4

5. 3. MgCL2 2μl is used per sample. Functions as a cofactor of Taq DNA polymerase by providing magnesium ions. Increase polymerase activity of Taq polymerase. 5

6. 4. dNTPs dNTP  stands for deoxyribonucleotide triphosphate . 0.2 μl is used per sample. There are four different  dNTPs . The dNTPs serve as the building blocks for the new strands being processed during PCR.  6

7. 5. PRIMERS PCR primers  are short fragments of single stranded DNA (15-30 nucleotides in length) that are complementary to DNA sequences that flank the target region of interest. The   purpose  of  PCR primers  is to provide a “free” 3'-OH group to which the DNA polymerase can add dNTPs . They are initiators of polymerization . Two types : Forward and Reverse Primers 1 μl Forward Primer and 1μl R everse primer is used in PCR per sample. 7

8. 6. Taq Polymerase Obtained from a bacterium Thermus aquaticus . High temperature stability. Performs the function of elongation of DNA strands . High rate of dNTP incorporation.   No proof reading ability. 0.4 μl is used per sample. 8

9. 7. TARGET DNA SEQUENCE The Target DNA sequence which need to be amplified . It is also called amplicon . 2 μl is used per sample. 9

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