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 cdna libraries, A cDNA library is made by using the reverse transcr...

A cDNA library is made by using the reverse transcriptase of a retrovirus  to synthesize cDNA (complementary  DNA)  copies  of the total mRNA from a cell (or perhaps a subfraction

Ask, what are the names of karyotypes?

what are the names of karyotypes?

Realization of p-d polarity of limb, Realization of P-D Polarity of Limb ...

Realization of P-D Polarity of Limb Realization of P-D polarity of limb expressed in a specified proximo-distal sequence of differentiation of limb pans is controlled through

Theoretical foundation of halsted reitan battery, Theoretical Foundation of...

Theoretical Foundation of Halsted Reitan battery There are really two theoretical bases for the Halsted Reitan battery, one contained in brain and intelligence and related writ

Why the recipient important in transfusions, Why is the determination of th...

Why is the determination of the blood types of the donor and of the recipient important in transfusions? Red blood cells have dissimilar antigens in the outer surface of their

Special uses of biogas, S p e c i a l Uses Biogas Biogas is an im...

S p e c i a l Uses Biogas Biogas is an important renewable energy resource for rural areas. Generation of bio-gas from dung and urine is a very simple and cost-effective

In what ways does over gazing lead to soil erosion?, Ask In what ways does ...

Ask In what ways does over gazing lead to soil erosion?

Difference between vegetative and generative cell , Difference between Vege...

Difference between Vegetative and Generative Cell The cytoplasm of the vegetative cell and the generative cell is distinctly different. The generative cell is transparent and

Sulphur cycle, Sulphur is one of important of proteins vitamins and hormone...

Sulphur is one of important of proteins vitamins and hormones. It plays a very crucial role in health of both plants and animals. Most of earth sulphur is tied up in rocks and salt

Is there a larval stage in echinoderms, Q. Is there a larval stage in echin...

Q. Is there a larval stage in echinoderms? In echinoderms embryonic development is indirect, with ciliated larvae.

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