HYS CH6.5 Recombinant DNA and Biotechnology

Question 1

plasmid

Answer 1

circular double stranded DNA

Question 2

recombinant DNA

Answer 2

could use bacterial plasmid and human gene
isolate gene from the plasmid

Recombinant DNA technology comprises altering genetic material outside an organism to obtain enhanced and desired characteristics in living organisms or as their products. This technology involves the insertion of DNA fragments from a variety of sources, having a desirable gene sequence via appropriate vector

Question 3

transformation

Answer 3

introduces new genetic information into host so it can integrate the new DNA into its genome

Question 3

restriction enzymes

Answer 3

site specific that cleave specific areas of plasmid to leave sticky ends that later hybridize

Question 4

PCR

Answer 4

• uses primers (complementary) to bind to target with optimal high G-C content as additional 3H bonds are more stable
• heat to denature
• uses Taq bacterial DNA polymerase which doesnt denature like human one
• dNTPs (nucleotides)
• DNA polymerase

1. DNA denaturation seperates strands
2. hybridization primers anneal to complements, dNTPs extend with DNA polymerase in a thermocycler (at least 70 degrees celcius
3. replication: every n, 2^n DNA strands
4. annealing of daughter strands
   doubles amount of DNA each cycle

Question 5

gel electrophoresis

Answer 5

**how many bp is the DNA fragment? (SIZE)
**
moves fragments (smaller ones move faster) down a gel. all move forward bc DNA is neg and moves to + end

use a DNA ladder of varrying segments to compare our samples to

Question 6

how do we identify fragment of DNA from solution of varying length?

Answer 6

Southern blots use probes to identify presence and quantitity of DNA after its on a gel (is target gene in a sample)

nitrocellulous paper plots DNA
flourescent probe placed which binds to single strand fragments

primer only binds to specific sequence and band means we have the gene of interest

Question 7

inserting genetic material is called

Answer 7

subcloning

we want a plasmid what will knock in the gene

prep with a restriction site specific enzyme

tranform plasmid product with E coli and only the bacteria with gene should group with an antibiotic plate

confirms it can express it!

Question 8

gene therapy

Answer 8

curing genetic deficiencies by introducting a functional gene with a viral vector

Question 9

DNA sequencing

Answer 9

uses dideoxyriboses which terminate DNA bc they have no 3’ OH and fragments can be seperated by gel electrophoresis and sequence read by the cell

Question 10

hybridization

Answer 10

process of joining complementary base pairs

Question 11

cDNA libraries

Answer 11

cDNA libraries have small fragments of DNA using only exons of genes in a tissue use for recombinant proteins or gene therapy

Question 12

genomic libraries

Answer 12

large fragments of DNA, coding and noncoding regions of genome which make recombinant proteins for gene therapy