Recombinant DNA Technology And Biotechnology 11-21

Question 1

Cystic fibrosis is caused by a ________ mutation

Answer 1

Genetic

Question 2

Cystic fibrosis results in defective what

Answer 2

Cl ion channeling

Question 3

What causes defective Cl ion channeling in cystic fibrosis

Answer 3

Due to improper folding and assembly of CFTR-cystic fibrosis transmembrane conductance regulator-chloride transport protein prematurely degraded

Question 4

What is the most common lethal genetic mutation in caucasians?

Answer 4

Cystic fibrosis

Question 5

Clinical diagnostic hallmark of cystic fibrosis

Answer 5

Elevated chloride levels in sweat- specific to CF

Question 6

Cystic fibrosis affect on lungs

Answer 6

Lack of chloride secretion leads to dehydration of the mucus, mucus accumulates in the lungs, obstructs airway passages, predisposed to recurrent lung infections

Question 7

Other symptoms of cystic fibrosis

Answer 7

Also causes pancreatic enzyme insufficiency and poor nutrition among other problems

Question 8

What are most cases of cystic fibrosis due to?

Answer 8

A 3 nucleotide deletion in the CTFR gene that leads to a missing phenylalanine residue

Question 9

Detection and diagnosis of common CF mutation

Answer 9

Using ASO probes (allele specific oligo

Question 10

PCR and cystic fibrosis

Answer 10

PCR using primers specific to the region of the deletion-PCR product is only a portion of the gene
-other known mutations causing CF would require different analysis

Question 11

Sickle cell anemia and cystic fibrosis genetic diagnosis

Answer 11

Very specific mutations-single nucleotide change in sickle cell anemia and 3 nucleotide deletion in CF

USE ASO PROBES AND PCR

Question 12

What types of tests would you do for sickle cell anemia and cystic fibrosis

Answer 12

ASO probes and PCR

Question 13

PKU

Answer 13

All newborns are screened for PKU-blood test for elevated phenylalanine levels and low levels of tyrosine
-400 different known mutations

Question 14

Where can mutations occur in PKU

Answer 14

In any of the 13 exons

Question 15

What type of mutations are PKU

Answer 15

Majority are missense, some splice, nonsense, insertion, and deletion mutations

Question 16

How big are introns for PKU

Answer 16

HUGE

Question 17

To do a genetic screen for PKU we must have:

Answer 17

1. The DNA from several family members including an affected individual
2. An identifiable marker associated (linked) with the particular mutation in this family (an RFLP)

Question 18

What would the marker be for PKU?

Answer 18

Polymorphisms (nucleotide change that results in an added or lost restriction site)
-generally the marker is not actually the disease causing mutation

Question 19

Markers are at least _____ accurate in predicting the presence of a mutation

Answer 19

98%

Question 20

What happens when a marker is identified in PKU?

Answer 20

A genetic screen can be performed (determine if a fetus has it)

Question 21

If a couple has one affected child and two phenotypically normal children, both parents are ______ carriers.

Answer 21

Obligate. Neither parent expresses it

Question 22

Myotonic dystrophy

Answer 22

• trinucleotide repeat expansion disease in the 3’ non-coding region of a proline kinase gene
• no phosphatase, cant relax muscles

Question 23

What is the most common form of adult muscular dystrophy?

Answer 23

Myotonic dystrophy

Question 24

Because myotonic dystrophy is associated with repeat expansion, it will produce _____ when digested with a restriction enzyme on both sides of the expansion

Answer 24

RFLPs

Question 25

What happens to expansions as you go though generations?

Answer 25

One mutant allele increases in size in subsequent generations (correlates to severity of disease)

Question 26

____ can be used to diagnose triplet repeat expansions

Answer 26

PCR

Question 27

Why can you not use PCR for myotonic dystrophy?

Answer 27

The repeat can be so huge that PCR is difficult

Question 28

Current methodology for testing for HIV uses __________ for diagnosis of HIV exposure. Why?

Answer 28

• immunoassay
• after exposure to HIV the body produces antibodies to HIV, immunoassay use enzymes linked antibody that recognize HIV antibodies

Question 29

What two tests do you use to detect HIV?

Answer 29

1. ELISA

2. Western blot

Question 30

Why do we need to do western blot after ELISA for HIV?

Answer 30

ELSA is so sensitive that it can produce false positives.

Question 31

What do immunoassay using antibodies to HIV-antibody require

Answer 31

That the body has mounted an immune response to a detectable level- requires a certain incubation time (6 months)

Question 32

Can use ______ for HIV provirus to test immediately

Answer 32

PCR

Question 33

__________ is used to monitor viral load in HIV positive patients

Answer 33

Quantitative PCR

Question 34

What is tested in paternity testing?

Answer 34

VNTRs

Question 35

What test do you do for paternity testing

Answer 35

Usually done using PCR

Question 36

Legal accuracy of paternity tests

Answer 36

Differs in different states, paternity index is a measure of accuracy

Question 37

Why do paternity cases always require several different VNTRs to be used?

Answer 37

Different VNTRs have a different paternity index

Question 38

Testing forensics

Answer 38

Restriction digests, southern blot with probe for tandem repeats (VNTR)

Question 39

Pattern of ______ can be used as a molecular fingerprint in forensics

Answer 39

RFLPs (VNTRs)

-normally several restriction digests and several different probes would be tested

Question 40

What is a good method if the amount of DNA evidence is too small to do southern blotting? (Forensics)

Answer 40

PCR based approaches