Ch. 14 Molecular Genetic Analysis And Biotechnology Flashcards
Recombinant DNA technology
set of molecular techniques for locating, isolating, altering, and studying DNA segment. RECOMBINANT means combine DNA from two distinct sources.
restriction enzymes (restriction endonucleases
recognize and make double-stranded cuts in DNA at specific nucleotide sequences. these enzymes are produced naturally by bacteria and are used in defense against viruses.
cohesive ends
“sticky ends” they are complementary to each other and can spontaneously pair to connect fragments. a lot of restriction enzymes produce sticky ends.
engineered nucleases
genetically engineered more complex restriction enzymes. capable of making double stranded cuts to the DNA at any predetermined DNA sequence.
CRISPR RNAs
developed genome manipulation technique uses these. small RNAs found in bacteria and archaea. couple with nucleases that cut dna.
gel electrophoresis
separate DNA molecules bases on size and elecrtrical charge. uses porous gele agarose.
cloning vector
stable replicating DNA molecule into which a foreign DNA fragment can be inserted for introduction into a cell.
plasmid
typically a small circular DNA strand in the cytoplasm of a bacterium or protozoan. Plasmids are much used in the laboratory manipulation of genes.
Cosmids
plasmids that are packaged into empty viral protein coats and transferred to bacteria by viral infection. carry more than twice as much foreign DNA as can a phage vector.
Bacterial artificial chromosomes (BACs)
vectors originally constructed from the F plasmid. can hold very large fragments of DNA that can be as long as 300,000 bp.
expression vector
a foreign gene is typically inserted into an EXPRESSION VECTOR: which, in addition to the usual origin of replication unique restriction sites and selectable markers contains sequences required for transcription and translation in bacterial cells.
Polymerase chain reaction (PCR)
- Kary Mullis. allows DNA fragments to be amplified a billionfold in just a few hours.
How to carry out PCR
solution that includes target DNA, DNA polymerase, all four deoxyribonucleoside triphosphates (dNTPs - ste substrates for DNA polymerase) primers, and mg ions and other salts necessary for reaction to proceed. Taq polymerase can be used due to withstanding high temperatures.
DNA sequencing
A powerful molecular method for analyzing dna is a technique. determines the sequence of bases in dna.
ddNTP
in the sanger method a special nucleotide called dideoxyribonucleoside triphosphate is also used as a substrate. ddNTPs are incorporated into growing DNA strand. no more nucleotides can be added, thus they terminate DNA synthesis. LACK 3’ OH group.
