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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

intracellular vesicles are found in
Akshay Reply
how do i report widal slide results
Isaac Reply
how food can be used as a substrate for the growth of microorganisms
Ashi Reply
what is the importance of understanding chemistry in the field of microbiology and food technology?
Charmaine Reply
What is different between eukaryotes and prokcaryotic
Abia Reply
main difference is that eukaryotic cells possess membrane bound organelles
Iqra
prokaryotes are primitive organisms that doesn't contain membrane bound nucleus or any orgenelle while in eukaryotes membrane bound orgenelles and nucleus is present
Yashi
what s anatomy
jane Reply
its mean body structure, function, and systems
gazi
study of internal structure of living things
Falere
anatomy is the study structure of the made human
isir
anotomy is the actual study of body internally and externally Which include how itis made. for what and what is need of this
Iqra
la science anatomie c'est la science qui nous aide à étudier l'homme
Baraka
okay
Baraka
Ancestor are they real
Rapheal Reply
yes of course
David
yes please
Clark
50 50, depending on the accuracy of the clan records.
Vincent
okay
Baraka
Why protist is not a kingdom of Linnaeus 'S taxonomy?
Neha Reply
I dont know
Jeewraj
Linnaeus used Aristotle's criteria for dividing living organisms into kingdoms. Plants are immobile and insensitive, while animals are mobile and sensitive. the creatures he saw through the microscope were mobile, so he attributed them to animals.
Lad
name the different types of media use in lab to detect the micro organisms
Shehzadi Reply
The different types of media used in the lab to detect the microorganisms is known as cell culture..
Tean
solid media or broth is used to grow and detect microorganisms
Pohor
what is microbiology
Baraka
the study of large living organisms
Pratibha
the study of organisms which are micro in range
Vency
what is autoclaving?
Yashi
process for sterilization
Vency
is a machine used on the process of sterilisation
Clark
what is difference between hot air oven and autoclave as they both are used for sterilization ?
Vency
autoclave basically do moist heat sterilization while hot air oven do sterilization by dry heat.....
Yashi
what is the microbe
HUSSAIN
which method out of these two is best?
Yashi
why human have microbe
HUSSAIN
Hi
Sadam
hi to all
Sadam
depends on what to sterilize
Vency
microbes
Md
what is knowledge
HUSSAIN
well idea
Daniel
what are the importance of microbiologe to a nurse
Enny Reply
it enable a nurse to know the weight and height of a patient
Abotu
it helps the nurse in way as to give first aid to patient on which basis doctor will take diagonos
Iqra
disagree with both of you
Akhtar
ok
Abotu
Your Own idea
Abotu
it helps a nurse to be able to counsel a patient /client
Abotu
cell culture
Tean
Types of micro organisms
Jennifer Reply
Who discovered plant and animal cell
Jennifer
Robert Hooke
Martha
Father of microbiology
Jennifer Reply
Antonie van Leeuwenhoek
Satarupa
Why are mitochondria and chloroplasts unable to multiply outside of a host cell?
Iqra Reply
iqra mitochondria arises from the division of existing mitochondria and they are fused together. they move around inside the cell with the interactions of the cytoskeleton that's why mitochondria unable to multiply outside the host cells chloroplasts has its own separate DNA from the plant cell.
Rana
short not on medical microbiology
Massah Reply
Hy
Iqra
hope you fine
Arif
love you
Arif
Assalamualaikum
Suhaib
arai yeh micro kai sth kis ko love hua
Umer
icrobiology is the scientific study of these microorganisms. Microorganisms are those organisms that are too small to see with the naked eye and include things like bacteria, fungi, and viruses
Suhaib
iqra iqbal
Arif
What is gram syaining?
Arif
to differentiate between gram positive and gram negative bacty
Umer
True
Arif
where you are from
Arif
i m frm sopore..kashmir
Umer
i m frm baramulla .. Kashmir
Umer
and u
Umer
umer
Arif
and i m from afghanistan lovely country
Arif
yes
Umer
thanks
Arif
medical microbiology means study of microorganisms which are beneficial or harmful for the body. mostly which are studied under microscope by structure and shape appearance. e.g bacteria, viruses, bacteriophage, fungus hyphae and some blood and other body fluids parasites e.g plasmodium etc.
Rana
arif bhai gram staining in which we are given different dyes to the bacterial cell wall and cytoplasm and nucleus.e.g giemsa stain
Rana
Gram staining uses crystal violet to stain cell walls iodine as a mordant and a fuchsin or safranin counterstain to mark all bacteria
Umer
Gram staining uses crystal violet to stain cell walls iodine as a mordant and a fuchsin or safranin counterstain to mark all bacteria
Umer
Gram staining uses crystal violet to stain cell walls iodine as a mordant and a fuchsin or safranin counterstain to mark all bacteria
Umer
Omar baba crystal violet iodine 70% alcohol fuchicine or safranine after every step washing must be done
Rana
hello
Cristy
*Fuchsin stain
Cristy
*Safranin
Cristy
thanks
Ganesh
difference between epidermophyton trichophyton and microsporum
Ankita Reply
plz answer the question
Iqra
epidermophyton type of fungi causes superficial and cutaneous mycoses trichophyton is also fungi type including parasitic varieties causes dermatophytosis microsporum is also type of fungi causes dermatophytosis.
Rana
ok
Iqra
Is this app perfect for preparation of exam?
Ankita
Ankita I totally disagree with your respected opinion. it's just minor helpful
Rana
disagree by me ?
Ankita

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Source:  OpenStax, Microbiology. OpenStax CNX. Nov 01, 2016 Download for free at http://cnx.org/content/col12087/1.4
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