Enzymatic mutation detection method, Biology

Assignment Help:

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.

 


Related Discussions:- Enzymatic mutation detection method

What is standard operating procedure manual (sop), Question 1 What is s...

Question 1 What is standard operating procedure manual (SOP)? Do you think it is important to maintain SOPs in all the laboratories? List the advantages of maintaining SOPs in

Gas exchange, Gas Exchange Gill surface area must be large enough to p...

Gas Exchange Gill surface area must be large enough to provide adequate exchange of gases. Therefore, highly active fish have largest relative gill area. Figure compares highl

Single vessel disease, Single Vessel Disease (SVD) :  They do well on medi...

Single Vessel Disease (SVD) :  They do well on medical treatment or with angioplasty. However if proximal LAD is significantly blocked and LIMA can be used as a conduit, surgery g

Benthos - aquatic ecosystem, Benthos - Aquatic Ecosystem The benthos o...

Benthos - Aquatic Ecosystem The benthos or the benthic organisms are those found living in or on the bottom or benthic region of the water mass. They exhibit a variety of adap

Fresh water-types of water found on earth, Fresh Water Water, a univers...

Fresh Water Water, a universal solvent, invariably contains many soluble salts. In fresh water the total salt content remains under 15 percent. Different types of soluble salts

Explain the minimum requirement of nutritional needs, Explain the Minimum R...

Explain the Minimum Requirement of nutritional needs? Minimum nutrient requirement is defined as the 'lowest amount of the nutrient ji-om the diet that will prevent clinically

Bacteria, Bacteria The study of bacteria is called bacteriology. Bacte...

Bacteria The study of bacteria is called bacteriology. Bacteria are unicellular organisms and possess a distinct cell wall. They are included in kingdom Prokaryote. The bacter

Determine about the halstead''s method, Determine about the Halstead's meth...

Determine about the Halstead's method Reitan adopted Halstead's methods and various test procedures and with them established a laboratory at the University of Indiana. He supp

What happens at the molecular level, An enzyme isolated from a mutant bacte...

An enzyme isolated from a mutant bacterium grown at 20 degrees celsius works in a test tube at 20 degrees celsius but not at 37 degrees celsius( 37 degrees celsius is the temperatu

Indications for surgery-pericardial effusion, Pericardial Effusion:  Indica...

Pericardial Effusion:  Indications for Surgery: Pericardial effusion may be the result of peiicarditis due to infection, autoimmune disease or neoplasm. Non-inflammatoly diseases

Write Your Message!

Captcha
Free Assignment Quote

Assured A++ Grade

Get guaranteed satisfaction & time on delivery in every assignment order you paid with us! We ensure premium quality solution document along with free turntin report!

All rights reserved! Copyrights ©2019-2020 ExpertsMind IT Educational Pvt Ltd