Alkalose Gel Analysis
Alkalose Gel Analysis - Related Videos
05:06
Alkaline Single Cell Gel Electrophoresis: A Sensitive Technique for Quantitative Estimation of DNA Damage in Individual Cancer Cells Following Chemotherapy
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2025
In this video, we demonstrate an alkaline single-cell electrophoresis assay to quantify both single- and double-strand DNA breaks in doxorubicin-treated cells. Alkaline conditions disrupt the hydrogen bonds between DNA base pairs, resulting in the complete unwinding and separation of DNA strands. This allows differential migration of undamaged and damaged DNA fragments in agarose gel upon electrophoresis, revealing a comet-shaped structure of slowly migrating undamaged DNA forming the circular...
10:38
In Vivo Alkaline Comet Assay and Enzyme-modified Alkaline Comet Assay for Measuring DNA Strand Breaks and Oxidative DNA Damage in Rat Liver
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Cited by 17 •
2016
The alkaline comet assay measures DNA strand breaks in eukaryotic cells. By adding an Endonuclease III or human 8-oxoguanine-DNA-N-glycosylase digestion step, the assay can efficiently detect oxidative DNA damage. We describe methods for using these assays to detect DNA damage in rat liver.
04:59
Colorimetric Analysis of Alkaline Phosphatase Activity in S. aureus Biofilm
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Cited by 9 •
2019
In this manuscript, we established a high throughput method to quantify alkaline phosphatase activity in S. aureus biofilm culture from 96-well tissue culture...
11:29
Modified Terminal Restriction Fragment Analysis for Quantifying Telomere Length Using In-gel Hybridization
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Cited by 10 •
2017
In this article, a detailed protocol for quantifying telomere length using a modified terminal restriction fragment analysis is discussed that provides fast and efficient direct measurement of telomere length. This technique can be applied to a variety of cell sources of DNA for quantifying telomere length.
06:30
Gel Purification
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2023
Gel purification is used to recover DNA fragments after electrophoretic separation. DNA recovery from an agarose gel includes three basic steps: binding, washing and eluting from a silica column. DNA is believed to bind to silica in the presence of high salt via a salt bridge. Following binding, DNA is washed of impurities and eluted under low salt conditions disrupting this interaction. This video goes through a step-by-step, generalized procedure for cutting out a band from the gel, gel...
FAQs
High pH separates the two DNA strands, changing the molecular forms that enter the gel and making strand-related integrity differences observable. During electrophoresis, charged nucleic-acid molecules move toward the positive electrode, while fragment mobility depends mainly on size and conformation. Thus, the pattern reflects both molecular length and structural state.
Size and conformation influence migration in different but related ways. Fragment length affects how readily DNA moves through the agarose matrix, whereas conformation describes the molecule’s structural form and can alter its mobility. Considering both variables prevents band position from being interpreted as a simple size measurement when samples may contain damaged or structurally distinct DNA.
Band patterns provide a comparative readout of DNA condition rather than an isolated label for one event. Differences among samples can reveal increased fragmentation, reduced strand integrity, altered replication intermediates, or damage associated with a biological or chemical stress. Interpretation is strongest when the pattern is related to the experimental condition being examined.
The agarose matrix provides a medium through which separated nucleic-acid molecules migrate under an electric field. In the alkaline format, its high-pH environment works with strand separation to expose differences in DNA fragmentation, integrity, replication-related structures, and stress-associated damage. The matrix therefore supports both separation and subsequent pattern comparison.
The central setup combines an alkaline agarose matrix, high-pH conditions, and an electric field with a positive electrode toward which charged molecules migrate. After separation, researchers examine the resulting band pattern, considering fragment size and conformation when comparing samples. These features are essential for linking migration behavior to DNA molecular state.
In biology, the method helps investigators compare molecular changes among DNA samples exposed to different biological or chemical stresses. Its band patterns can indicate fragmentation, strand-integrity changes, replication intermediates, or damage. These observations support research on genome stability and DNA repair by connecting experimental conditions with changes in nucleic-acid structure.