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An Improved Modified Protocol for Silver Staining of Ds DNA in Agarose Gel.
Author(s):
1. Nuzhat Aisha Akram: Karachi Institute of Medical Sciences (KIMS) Karachi, Pakistan
2. Bushra Iftikhar: Combined Military Hospital Malir/National University of Medical Sciences (NUMS), Pakistan
3. Shoaib Ahmed: Karachi Institute of Medical Sciences (KIMS) Karachi, Pakistan
4. Fasiha Fatima: Karachi Institute of Medical Sciences (KIMS) Karachi, Pakistan
5. Tehmina Qamar: Karachi Institute of Medical Sciences (KIMS) Karachi, Pakistan
6. Shakeel R Farooqi: Karachi Institute of Medical Sciences (KIMS) Karachi, Pakistan
Abstract:
Objective: Present study was aimed to develop a reproducible, cheap and sensitive method for silver staining of double stranded DNA in agarose gel. Study Design: Experimental, repeated measure design. Place and Duration of Study: Department of Genetics, University of Karachi, Karachi. This experimental study was conducted, from Nov 2013 to Jan 2014. Material and Methods: The new method is the modification and improvement in the original method proposed in the literature. Samples of ds genomic DNA was run on a nondenaturing 1.5% agarose/0.5X TBE. After electrophoresis gel was fixed in 10% acetic acid and staining was performed using 1 gm % silver nitrate. DNA bands were developed using 1.5% NaOH. At each step shaking was done manually with a circular movement. The modified method was also compared with the ethidium bromide staining of the same samples of DNA. Results: The modified method was proved to be as sensitive as the ethidium bromide with the advantage of having long term conservation ability of the gel. The main advantage of the protocol is the consumption of far less concentrations of silver nitrate and sodium hydroxide and the avoidance of the use of sodium thiosulphate. Conclusion: This method was easily reproducible, sensitive, and cheap with improved conservation ability of gel. It also avoids use of hazardous, expensive and time consuming radioactive and fluorescent detection.
Page(s): 87-91
DOI: DOI not available
Published: Journal: Pakistan Armed Forces Medical Journal, Volume: 69, Issue: 1, Year: 2019
Keywords:
Silver staining protocol , dsDNA
References:
[1] MohantyAK,YadavML,ChoudharyS. Gel, Singapore 2017.Electrophoresis of Proteins and Nucleic Acids,In Protocols in Semen Biology (Comparing Assays) 233 -46
[2] LiuW,LiR,AyalewH,XiaY,BaiG,YanG, 2017.Development of a simple and effective silver staining protocol for detection of DNA fragments,Electrophoresis 38 1175 -8
[3] HuangL,DengX,LiR,XiaY,BaiG,SiddiqueKH, 2018.A fast silver staining protocol enabling simple and efficient detection of SSR markers using a non-denaturing polyacrylamide gel,J Vis Exp 20 1 -9
[4] LiangQ,WenD,XieJ,ShiS,LiuLL,WeiYZ, 2014.A rapid and effective method for silver staining of PCR products separated in polyacrylamide gels,Electrophoresis 35 2520 -3
[5] ParedesAJ,PalmaNT,ValdésAHM,BarrigaA,BabulJ, 2017.New visible and selective DNA staining method in gels with tetrazolium salts,Anal Biochem 517 31 -5
[6] Kumar , M,KimSR,PC , Pareek,A, 2015.Simple and efficient way to detect small polymorphic bands in plants,Genomics Data 5 218 -22
[7] BassamBJ,GresshoffPM, 2007.Silver staining DNA in polyacrylamide gels,Nature Protocols 2 2649 -54
[8] TangW,ZhouH,LiW. Silver, 2015.and cyanine staining of oligonucleotides in polyacrylamide gel,PloS one 10 e0144422 -
[9] BenbouzaH,JacqueminJM,BaudoinJP,MergeaiG, 2006.Optimization of a reliable, fast, cheap and sensitive silver staining method to detect SSR markers in polyacrylamide gels,Biotechnol Agron Soc Environ 10 77 -81
[10] QiuS,ChenJ,LinS,LinX., 2012.A comparison of silver staining protocols for detecting DNA in polyester backed polyacrylamide gel,Braz J Microbiol 43 649 -52
[11] SwitzerRC,MerrilCR,ShifrinS., 1979.A highly sensitive silver stain for detecting proteins and peptides in polyacrylamide gels,Anal Biochem 98 231 -7
[12] OakleyBR,KirschDR,MorrisRN, 1980.A simplified ultrasensitive silver stain for detecting proteins in polyacryamide gels,Anal Biochem 105 361 -3
[13] SommersvilleLL,WangK.,Biochem, 1981.The ultrasensitive silver protein stain also detect nanograms of nucleic acids,Biochem Biophys Res Commun 102 53 -8
[14] GottliebM,ChavkoM., 1987.Silver staining of native and denatured eucaryotic DNA in agarose gels,Anal Biochem 165 33 -7
[15] WilloughbyEW,LambertA, 1983.A sensitive silver stain for proteins in agarose gels,Anal Biochem 130 353 -8
[16] PeatsS., 1984.Quantitation of protein and DNA in silver stained agarose gels,Anal Biochem 140 178 -82
[17] LasneF,BenzeraraO,LasneY, 1983.,Anal Biochem 132 338 -431
[18] BrownTA, 2002.Essential Molecular Biology: A Practical Approach,2nd Edition 1 102 -03
[19] LiW,GaoF,ZhangXR,TangWZ,ZhangHT, 2006.Detection of known thalassemia point mutations by snapback single strand conformation polymorphism: The feasibility analysis,Clin Biochem 39 833 -42
[20] PrietoC,LeonardelliRI,ZalazarFE, 1997.A procedure of silver staining for nucleic acids in agarose gels without pretreatment or drying steps,Anal Biochem 252 15 -8
[21] ZalazarFE,BaroniMR,AmatoD,GianniR,MayralC,OchotecoC, 2001.Modifieddd silver staining for RNA and DNA in agarose gels,Anal Biochem 291 299 -300
[22] SambrookJ,FritschEF,ManiatisT. Molecular,Cloning, 1989.A laboratory manual, -
[23] HempelmannE,KraftsK., 2017.The mechanism of silver staining of proteins separated by SDS polyacrylamide gel electrophoresis,Biotech Histochem 92 1 -7
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