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Hi there
Some basic concepts..
#1..What is the difference between heterozygous and homozygous?
#2..What exactly does restriction fragments length polymorphysim mean?Does that mean that the normal genes associated with restriction fragments which are normally present in everyone makes a total length smaller than when the mutation occurs?Does that add the restriction fragments and the gene area?
#3...What is ELISA?In southern blot why it is done as a first step?
#4...In PCR why doesnot it need blotting techniques.If it is just an amplification technique,How can it detect genetic defects?
I am really helpless in these questions.If someone is kind enough to explain,I will be greatly obliged.
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| vermdocamit
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Heterozygous-- for one "thing" in an
individual two different gene type on the chromosome "Gene A + Gene a"
Gene A-- from mama
Gene a--- from papa
or vice versa
Homozygous---- for one "thing" in an
individual two different gene type on the chromosome "Gene A + Gene A"
Gene A --- from mama
Gene A---- from papa
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| vermdocamit
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RFLP ( restriction fragment length polymorphism)
Restriction enzymes "cut " the Nucleic acid----> "Fragments" of Nucleic acid
Suppose one fragment is------ "Fragment A"
Present on ------------ "Location 1" on a Blot.
If Nucleic acid of all the other individuals is found to have "Fragment A" on "Location 1"----- Monomorphism--representing same form.
To say that it is monomorphic is to say that it does not vary in form among the individuals of a population.
If Nucleic acid of other individuals is found to have "Fragment A" on "Location 1", "Location 2" etc on the blot----- Polymorphism
This is RFLP representing different forms of , say a gene in different individuals.
To say that it is Polymorphic is to say that Fragment though representing same thing is variable in form.
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| vermdocamit
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ELISA first step in Southern Blot ? :? never heard ( may be I am wrong)
I thought it was like this-
Digest the DNA( Restriction enzyme)----> place on Agarose---> denature
-->Electrophoresis ---> seperate bands---> Blot on Membrane-------------
-------->prehybridise ( to block non specific sites)---> specific probing--->
---->Detect
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| DR_K
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ELISA is "enzyme-linked immunosorbent assay" i did know that it is used in the first step of blotting but this methid is used to measure quntities of anigen and antibodies(probably proteins in this case) fr example u wanna measure ammount of some"X"anti body what u 'll do that u fixed known antigens &then put those anti bodies they can attached to each other and after thatu take a 3rd party fr instance'Y' antibodies which r labelled with enzyme these get attached to X antibodies(as they r designed fr this) and enzyme activity is measured by adding a substrate to that enzyme , the amount of antibody'Y' bound propotional to enzyme activity. i know the procedure but if u got to know how this is used in southern blotting plz let me know.
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| DR_K
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if a person has both the allele on a chromosome fr a particular disease he is homozygously effected fr that particular disease and if only one allele is mutant and the other normal he is heterozygously effected. simply
'AA' is homozygous and 'Aa' is hetrozygous
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| vermdocamit
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Blotting --- helps in detection
PCR is used so that we have something to blot !!
I mean i f a gene or nucleic acid we wish to detect is in small quantities you will need PCR
Hence PCR is used to amplify so that we have something we can blot. :lol:
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| beta blocker
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Guys you are great.You just cleared all my concepts.I am sorry if I am wrong to say that ELISA is the first step in southern blot.In kaplan biochemistry,ther is a flow sheet that has put elisa as first step for southern blotting.But I am weak in my concepts of biochem,so I dont know for sure.Thanx for the help all of you 
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| DR_K
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hey no probs NONONONO u were not wrong ab ELISA, i didnt know this and thank u so much fr drawing my attention towards its first step [/quote]
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