METHYLENE BLUE AS ALTERNATIVE DNA STAINING IN ELECTROPHORESIS

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Fusvita Merdeka wati
Ira Gustira Rahayu
Yogi Khoirul Abror

Abstract

Background: Electrophoresis is a technique for separating DNA molecules based on their size by using an electric charge. To make electrophoresis fast, precise, effective, ethidium bromide (EtBr) is used as an intercalator dye. However, EtBr is mutagenic and carcinogenic and therefore has significant disadvantages both from an environmental and human health perspective. To avoid the disadvantages of EtBr, scientists have started looking for alternative nucleic acid dyes. One of them is the use of fluorescent dyes which have high speed and sensitivity for DNA visualization.


Method: The type of research conducted is experimental research. The design of this study was made with variations in agarose concentration (1%, 1.5% and 2%), methylene blue concentration (0.00625%, 0.0125% and 0.025%), agarose gel contact time from electrophoresis with Methylene Blue and long destaining time (10, 20 and 30 minutes) and electrophoresis conditions in the form of length of time (1 hour, 1.5 hours and 2 hours) and voltage variations (100 volts and 150 volts) of running electrophoresis.


Result: the best appearance of the Sars CoV 2 DNA band at 100 bp and 250 bp using a concentration of 0.025% methylene blue, 1.5 % agarose concentration, 20 minutes of staining and 30 minutes of destaining. The electrophoresis process takes place with a voltage of 100 volts for 60 minutes.


Conclusion: The best appearance of the DNA band using a concentration of 0.025% methylene blue, Therefore, it is necessary to conduct further research regarding the sensitivity of Methylene Blue


Keywords: Methylene Blue, Alternative Staining, Electrophoresis


 


 


Background: Electrophoresis is a technique for separating DNA molecules based on their size by using an electric charge. To make electrophoresis fast, precise, effective, ethidium bromide (EtBr) is used as an intercalator dye. However, EtBr is mutagenic and carcinogenic and therefore has significant disadvantages both from an environmental and human health perspective. To avoid the disadvantages of EtBr, scientists have started looking for alternative nucleic acid dyes. One of them is the use of fluorescent dyes which have high speed and sensitivity for DNA visualization.


Method: The type of research conducted is experimental research. The design of this study was made with variations in agarose concentration (1%, 1.5% and 2%), methylene blue concentration (0.00625%, 0.0125% and 0.025%), agarose gel contact time from electrophoresis with Methylene Blue and long destaining time (10, 20 and 30 minutes) and electrophoresis conditions in the form of length of time (1 hour, 1.5 hours and 2 hours) and voltage variations (100 volts and 150 volts) of running electrophoresis.


Result: the best appearance of the Sars CoV 2 DNA band at 100 bp and 250 bp using a concentration of 0.025% methylene blue, 1.5 % agarose concentration, 20 minutes of staining and 30 minutes of destaining. The electrophoresis process takes place with a voltage of 100 volts for 60 minutes.


Conclusion: The best appearance of the DNA band using a concentration of 0.025% methylene blue, Therefore, it is necessary to conduct further research regarding the sensitivity of Methylene Blue


Keywords: Methylene Blue, Alternative Staining, Electrophoresis


 


 Background: Electrophoresis is a technique for separating DNA molecules based on their size by using an electric charge. To make electrophoresis fast, precise, effective, ethidium bromide (EtBr) is used as an intercalator dye. However, EtBr is mutagenic and carcinogenic and therefore has significant disadvantages both from an environmental and human health perspective. To avoid the disadvantages of EtBr, scientists have started looking for alternative nucleic acid dyes. One of them is the use of fluorescent dyes which have high speed and sensitivity for DNA visualization.
Method: The type of research conducted is experimental research. The design of this study was made with variations in agarose concentration (1%, 1.5% and 2%), methylene blue concentration (0.00625%, 0.0125% and 0.025%), agarose gel contact time from electrophoresis with Methylene Blue and long destaining time (10, 20 and 30 minutes) and electrophoresis conditions in the form of length of time (1 hour, 1.5 hours and 2 hours) and voltage variations (100 volts and 150 volts) of running electrophoresis.
Result: the best appearance of the Sars CoV 2 DNA band at 100 bp and 250 bp using a concentration of 0.025% methylene blue, 1.5 % agarose concentration, 20 minutes of staining and 30 minutes of destaining. The electrophoresis process takes place with a voltage of 100 volts for 60 minutes.
Conclusion: The best appearance of the DNA band using a concentration of 0.025% methylene blue, Therefore, it is necessary to conduct further research regarding the sensitivity of Methylene Blue
Keywords: Methylene Blue, Alternative Staining, Electrophoresis


 


 


 


 


 


 

References

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