Chapter 14 Flashcards
Recombinant DNA Technology
Set of molecular techniques for locating, isolating, altering, combining, and studying DNA segments (Genetic Engineering)
Joining of two or more different DNA molecules in vitro
Restriction Enzymes
How are they produced?
What can be added?
Recognizing and cutting DNA at specific nucleotide sequences.
Naturally produced by bacteria to defend against viruses.
Adding methyl groups to the recognition sequence to protect itself from being digested by its own enzyme bacteria.
What are cohesive ends and blunt ends?
Cohesive ends are fragments with short, single-stranded overhanging end.
Blunt ends: even-length ends from both single strands
What is gel electrophoresis?
How can DNA fragments be visualized?
Separates molecules (like DNA) based on size and electrical charge. These molecules are visualized by using a dye or adding chemical labels to the DNA.
When a gel is stained with a DNA-binding dye and placed under UV light, the DNA fragments will glow, allowing us to see the DNA present at different locations along the length of the gel
What is dideoxy sequencing?
Similar to PCR
DNA Polymerase
Template
Primer (only 1)
dNTPs
Dideoxyribonucleoside triphosphate (ddNTP) lacks a 3 -OH group, which terminates DNA synthesis
What is next-generation sequencing?
Sequencing millions of fragments simultaneously per run
Illumina sequencing, pyrosequencing
What are expression vectors?
A vector which is designed to allow expression (transcription and translation) of the inserted section of DNA. The vector carries a promoter (normally inducible) on one side of the cloning site, and a transcription terminator on the other side.
What is biotechnology?
What are the five biotechnology harnesses the power of molecular genetics has?
The use of techniques in recombinant DNA technology to develop new products. Use of living organisms for production of valuable products
- Pharmaceutical Products
- Specialized Bacteria
- Use in agriculture
- Genetic Testing
- Gene Therapy
What does the Polymerase chain reaction (PCR) provide?
Provides a way to amplify a single copy of
a gene or DNA sequence to trillions of
copies
What are the four essential ingredients for PCR?
- DNA polymerase (Taq)
- Template DNA (double-stranded)
- Free nucleotides
- Short oligonucleotide primers (15-50 nucleotides in length) of known sequence that have been chemically synthesized.
What are designing primers in PCR?
Forward and reverse primers. The forward and reverse primers are oriented on opposite strands of the DNA
What are thermocyclers in PCR?
Instruments used to amplify DNA and RNA samples by the polymerase chain reaction.
What are the three temperature variations in PCR reactions?
- Melting or denaturing temperature (95 C)
- Annealing temperatures (55 C)
- Polymerization temperature (72 C)
MAP
Why use PCR?
DNA fingerprinting, detection of bacteria or viruses (particularly AIDS), and diagnosis of genetic disorders.
Genotype detection and mapping, detection of infectious diseases. A way of amplifying a piece of DNA and create trillions of copies.
What is gene cloning?
amplifying a specific piece of DNA via an organism (like a bacterial cell). Uses living cells
– Also allows alter DNA sequence and/or alter gene expression
