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Due to its negatively charged backbone, DNA is strongly attracted to a positive electrode. In agarose gel electrophoresis, the gel is oriented horizontally in a buffer solution. Samples are loaded into sample wells on the side of the gel closest to the negative electrode, then drawn through the molecular sieve of the agarose matrix toward the positive electrode. The agarose matrix impedes the movement of larger molecules through the gel, whereas smaller molecules pass through more readily. Thus, the distance of migration is inversely correlated to the size of the DNA fragment, with smaller fragments traveling a longer distance through the gel. Sizes of DNA fragments within a sample can be estimated by comparison to fragments of known size in a DNA ladder also run on the same gel. To separate very large DNA fragments, such as chromosomes or viral genomes, agarose gel electrophoresis can be modified by periodically alternating the orientation of the electric field during pulsed-field gel electrophoresis (PFGE) . In PFGE , smaller fragments can reorient themselves and migrate slightly faster than larger fragments and this technique can thus serve to separate very large fragments that would otherwise travel together during standard agarose gel electrophoresis. In any of these electrophoresis techniques, the locations of the DNA or RNA fragments in the gel can be detected by various methods. One common method is adding ethidium bromide , a stain that inserts into the nucleic acids at non-specific locations and can be visualized when exposed to ultraviolet light. Other stains that are safer than ethidium bromide, a potential carcinogen, are now available.

a) A diagram of the process of agarose gel electrophoresis. 1 – An agarose and buffer solution is heated and poured into a form. This result in a rectangular block with indents along one end labeled “S=sample wells”. 2 – When cooled, the agarose gel block contains small wells (S) where the sample will be place. 3 – Each sample is added to a separate well. Then the agarose gel is placed in a chamber that generates a charge across the gel. The samples are added using micropipettes. 4 – The solution within the chamber conducts the electric current generated by the chamber. The side nearest the sample well will have a negative charge; the other side will have a positive charge. 5 – DNA has a negative charge and will be drawn to the positive pole of the gel. Smaller DNA molecules will be able to travel faster through the matrix of the gel. 6 – One well will contain a DNA ladder, which has fragments of known size. This ladder is used to identify the sizes of the bands in the sample. The ladder looks like many bands in the gel; from top to bottom the sizes of the bands are – 2000 bp, 15000 bp, 1000 bp, 750 bp, 500 bp, 250 bp. The other lanes have a few bands of various sizes.
(a) The process of agarose gel electrophoresis. (b) A researcher loading samples into a gel. (c) This photograph shows a completed electrophoresis run on an agarose gel. The DNA ladder is located in lanes 1 and 9. Seven samples are located in lanes 2 through 8. The gel was stained with ethidium bromide and photographed under ultraviolet light. (credit a: modification of work by Magnus Manske; credit b: modification of work by U.S. Department of Agriculture; credit c: modification of work by James Jacob)

Restriction fragment length polymorphism (rflp) analysis

Restriction enzyme recognition sites are short (only a few nucleotides long), sequence-specific palindromes, and may be found throughout the genome. Thus, differences in DNA sequences in the genomes of individuals will lead to differences in distribution of restriction-enzyme recognition sites that can be visualized as distinct banding patterns on a gel after agarose gel electrophoresis. Restriction fragment length polymorphism (RFLP) analysis compares DNA banding patterns of different DNA samples after restriction digestion ( [link] ).

RFLP analysis has many practical applications in both medicine and forensic science . For example, epidemiologists use RFLP analysis to track and identify the source of specific microorganisms implicated in outbreaks of food poisoning or certain infectious diseases. RFLP analysis can also be used on human DNA to determine inheritance patterns of chromosomes with variant genes, including those associated with heritable diseases or to establish paternity .

Questions & Answers

Explain about enzyme transportation
Shahla Reply
Enzyme transportation
what is the infectious disease process
Patience Reply
what are differences between endotoxins and exotoxins
sabote Reply
endo toxins work in the nuceus. i think
tell me if im right tho
Exotoxins are toxic substances secreted by bacteria and released outside the cell. Both gram positive and gram negative bacteria can produce and secrete exotoxins. Whereas Endotoxins are bacterial toxins consisting of lipids that are located within a cell. Only lysed gram negatives.
Remebr the Lipid A portion of LPS is what's toxic.
oh yeah. thanks
Your welcome :)
How did you learn this?
For me personally the best book is 'microbiology made ridiculously simple'
I got my basics from there and slowly added information from other sources.
thats cool! yeah i like microbiology too! especially the molecular proteins theyre sooooooooooo cool!
what are the prokaryotic
Lungu Reply
prokaraytotic is a unicellular organizm that lacks membrane bound nucleus
and whats eukaryotic
eukaryotic cell are cell which contain anuclues and organells
eukaryotes are the cells that have organells which are protected by membranes
eukaryotic is are multicellular organisms which are open nucleus.
Explain on the Francisco reddi did to prove the theory of spontaneous generation
Diana Reply
what is parasite
Abdirizack Reply
parasite are organisms feeds on a host for food and survival.e.g round worm for animal, Dodder for plant parasites.
parasite are organisms that feeds on a host for food and survival.e.g round worm for animal and mistletoe for plant parasites.
parasite are organisms that feed on their host
designing of aseptic area
Aashish Reply
I don't know
what is rickettsia
what is microbiology
Is the science that works with microorganisms.
richettsa is small microorganisms that cause disease in human like typhus; they are like viruses that can grow only inside living cells, they're transmitted by mites, ticks or lices.
what is plasmid?
plasmid is extra chromosomal body present in bacteria...which have additional genetic functions example... antibiotic resistance genes....etc etc
state the theory of spontaneous generation of micro oranisms and germ theory of disease
what are the advantages of high note numerical aperture
Genius Reply
list if non flagellated pritozoa
Mepung Reply
Can someone that's understanding of the Kreb Cycle please explain & breakdown it down to me in the simplest way without giving me the dictionary version or Google version. Basically in there own words of knowledge....!
please can someone help explain positive and negative feedback in simple term
negative feedback is the arresting of reaction or reverse of the reaction according to the response and postive feed back is the direct response without reversing or arresting a reaction.
pls can someone explained Kinney stone in memorising in shot time
what tyoebif microorganism will be killed by antibiotic trwatmeant
Mary Reply
I don't really don't understand aerobic respiration anaerobic respiration and fermentation. Can someone explain & breakdown this in the simplest way please.
Anaerobic Respiration in which foodstuffs are partially oxidized, with the release of chemical energy, in a process not involving atmospheric oxygen, such as alcoholic fermentation, in which one of the end products is ethanol.
aerobic respiration A type of respiration in which foodstuffs are completely oxidized to carbon dioxide and water, with the release of chemical energy, in a process requiring atmospheric oxygen.
please can someone explain what pseudomonas species and biofilm is?
Fermentation is the growth of cells or microorganisms in bioreactors (fermenters) to synthesize special products. Fermentation in biochemistry refers to the biodegradation of carbon compounds by cells or organisms under anaerobic (lack of oxygen) conditions.
Please under which conditions does pathogens become established in the human tissues and can cause diseases
please the life cycle of plasmodium parasite
who is John Needham
Mary Reply
John Needham is one of the researcher in microbiology. He also experimented when scientists did not believe animals could arise spontaneously ,but did believe microbes could.
okay is he late
by the way what are the list of courses offered by a newly admitted student for microbiology
Needham's experiments with beef gravy and infusions of plants material reinforced this idea.
what drugs are given to a person with Otis nerve problem(ear problem)
Gum Reply
good morning to you all.
Muhammad Reply
Dr.A.K.S.P.G. college Akbarpur Ambedkar Nagar
Shailesh Vishwakarma BSc 1st year

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