L.7 Recombinant DNA & Biotechnology Flashcards
What is Recombinant DNA?
DNA made from two different DNA’s
What is DNA Cloning?
DNA cloning introduces a fragment of DNA into a Vector Plasmid. A restriction enzyme cuts both the plasmid and the fragment, both are left with “sticky ends”.
When both bind, the transformed plasmids are introduced to bacterial cell to mass reproduce.
What 3 things must a vector contain?
- Origin of replication
- Fragment of interest
- Antibiotic resistance
DNA Libraries
Genomic Library vs cDNA Libraries
Genomic Libraries contain a full organism genome, both coding and non-coding, cannot be used to make recombinant proteins or gene therapy
cDNA Libraries are expression libraries only ccontain exons, they can be used to make recombinant proteins and for gene therapy
What is Hybridization?
it is the joining of complementary base pair sequences
What is and how does the PCR work?
Polymerase Chain Reaction
Using hybridization
To amplify a segment of DNA using PCR, the sample is first heated so the DNA denatures, or separates into two pieces of single-stranded DNA. Next, an enzyme called “Taq polymerase” synthesizes - builds - two new strands of DNA, using the original strands as templates. This process results in the duplication of the original DNA, with each of the new molecules containing one old and one new strand of DNA. Then each of these strands can be used to create two new copies, and so on, and so on. The cycle of denaturing and synthesizing new DNA is repeated as many as 30 or 40 times, leading to more than one billion exact copies of the original DNA segment.
What kind of gel is used to separate DNA
Agarose gel electrophoresis
Explain Southern Blotting
Southern blotting can be used to detect the presence and quantity of various DNA strands in a sample. After electrophoresis, the sample is transferred to a membrane that can be probed with single-stranded DNA molecules to look for a sequence of interest.
This probe can be tagged
What are dideoxyribonucleotides?
In DNA sequencing ddDNA terminate the DNA chain because they lack the OH at carbon 3
What is Gene Therapy used for?
Gene therapy is a method of curing genetic deciciencies by introducing a functional gene with a viral vector.
What is the difference between
transgenic mice and knockout mice?
Trangenic mice
Integratinf gene of interest into the germ line or embryonic stem cells of a developing mouse (chimeras)
Knockout mice
Created by deleting a gene of interest
