Enzymatic mutation detection method, Biology

This technique utilizes an enzyme resolvase, endo vii, cloned from the bacteriophage t4. This enzyme has high specificity to find deletions, insertions, and base substitutions mutations or any mismatches. First the pcr is utilized to intensify the normal and mutant allele of targeted sequence, these pcr primers are labeled with fam(6-carboxyfluoroscein amidite) which is a blue fluorescent dye and reverse primer with tet(4,7,2,7-tetra chloro-6-carboxy fluoroscein) amidite which is a green fluorescent dye.

Denaturation and renaturation of normal and mutant allele in a mixture is made, consequently mismatch heteroduplexes will form about 50 % of the time, each and every base pair creates two mismatch. The endo vii enzymes then scan the double stranded dna until it finds structural distortion, either a bubble made by single base pair mutations or a hetero duplex loop created by hybridization of a wild type allele with a mutant allele having an insertion or deletion. The enzyme cleaves with in 6 base pair on 3"side of mutation creating two smaller fragments, one blue and the other one green. The dna sequence is analyzed on automated dna sequence and mobility of every fragment is analyzed.

 

Posted Date: 3/6/2013 6:01:05 AM | Location : United States







Related Discussions:- Enzymatic mutation detection method, Assignment Help, Ask Question on Enzymatic mutation detection method, Get Answer, Expert's Help, Enzymatic mutation detection method Discussions

Write discussion on Enzymatic mutation detection method
Your posts are moderated
Related Questions
A decrease in parasympathetic discharge to the heart leads to A. an increase in the conductance of F-channels in SA node cells. B. a decrease in the conductance of potassium

Q. Qualitative and Quantitative Analysis of Colour? Primarily, two types of analysis are carried out for food colours - qualitative and quantitative. The qualitative analysi

Q. Explain about Treatment and Management of Dyslipidemia? Maintenance of blood lipids within the normal range is the most essential objective to prevent the onset/progression

Animal and Plant Cells The internal structural organisation and information related with hereditary characters and metabolic pathways are similar in all eucaryotic cells whethe

keel is the characteristic feature of flower of 1.tomato 2. tulip 3.ubdugifera 4.aloe

What are the types of muscle tissues? What are the morphological features that differentiate those types? There are three kinds of muscle tissue: the skeletal striated muscle t

Structural isomers: Select one: a. Have the same molecular weight b. Have the same connectivity c. Are mirror images d. All of the above e. None of the above

Explain Micafungin sodium Micafungin sodium (Mycamine - Astellas), the second echinocandin antifungal1 to become available in the US, has been approved by the FDA for intraveno

How does the placement of veins help to carry out their function of transporting materials to and from the leaf ?

Structure and Fate of Endosperm The cells of endosperm are usually thin-walled, large isodiametric and devoid of pits and store large amount of food materials. The starch alo