Chapter 20 Flashcards
recombinant DNA
nucleotide sequences from 2 different sources, often two species, are combined in vitro into the same DNA molecule
genetic engineering
the direct manipulation of genes for pratical purposes
biotechnology
the manipulation of organisms or their genetic components to make useful products
DNA cloning
well defined segments of DNA in identical copies
plasmids
small circular DNA molecules that replicate separately from the bacterial chromosome
why are cloned genes useful?
to make copies of a particular gene and producing a protein product
gene cloning
involves using bacteria to make multiple copies of a gene
- foreign DNA is inserted into plasmid, and then that plasmid is inserted into a bacterial cell
- reproduction in the bacterial cell results in cloning of the plasmid including the foreign DNA
- this results in the production of multiple copies of a single gene
restriction enzymes
cut DNA molecules at specific DNA sequences called restriction sites
restriction fragments
when a restriction enzyme makes many cuts this creates restriction fragments
sticky ends
most useful restriction enzymes cut DNA in a staggered way making sticky ends
DNA ligase
an enzyme that seals the bonds between restriction fragments
cloning vector
the original plasmid
-can carry foreign DNA in a host cell and replicate there
genomic library
the collection of recombinant vector clones produced by cloning DNA fragments from an entire genome
bacterial artificial chromosome (BAC)
large plasmid that has been trimmed down and can carry a large DNA insert
complementary DNA (cDNA)
made by cloning DNA made in vitro by reverse transcription of all the nRNA produced by a particular cell
nucleic acid probe
a way to identify a gene of interest… has a sequence complementary to the gene being seeked..this process is called nucleic acid hybridization
cloned genes can be expressed as protein in (bacteria or eukaryotic cells?)
BOTH
expression vector
a cloning vector that contains a highly active bacterial promoter
electroporation
applying brief electrical pulse to create temporary holes in the plasma membranes
-a method of introducing recombinant DNA into eukaryotic cells
polymerase chain reaction (PCR)
can produce many copies of a specific target segment of DNA
-a 3 step cycle (heating, cooling, and replication) brings about a chain reaction that produces an exponentially growing population of identical DNA molecules
Taq polymerase
a PCR that is very useful because it can work in very extreme temperatures and still produce copies of a specific target segment
what does cloning allow researchers to do (3)
- compare genes and alleles between individuals
- locate gene expression in a body
- determine the role of a gene in an organism
gel electrophoresis
- analyzing and comparing genomes
- separates nucleic acids by size, electrical charge (goes from negative to positive)
polymorphisms
variations in DNA sequence
RFLPs (restriction fragment length polymorphisms)
sequence changes that alter restriction sites
-a variation in the length of restriction fragments produced by a given restriction enzyme in a sample of DNA.
southern blotting
combines gel electrophoresis and DNA fragments with nucleic acid hybridization
what is used to compare mRNA from different developmental stages
- northern blotting
2. reverse transcriptase- polymerase chain reaction
northern blotting
an adaptation of the Southern blot procedure used to detect specific sequences of RNA by hybridization with complementary DNA.
reverse transcriptase polymerase chain (RT-PCR)
quicker and more sensitive because it requires less mRNA than Northern blotting
- added to mRNA to make cDNA, which serves as a template for PCR amplification of the gene of interest
- products are run on a gel and the mRNA of interest idnetified
situ hybridization
uses flourescent dyes attached to probes to identify the location of a specific mRNAs in place in the intact organism
DNA microarray assays
compare patterns of gene expression in different tissues, at different times, or under different conditions
in vitro mutagenesis
use this, mutations are introduced into a clone gene, altering or destroying its function
what are ways to determine gene function
- disable gene and observe consequences
- mutations with vitro mutagenesis then when mutated gene is returned to cell, the normal genes function might be determined by examining the mutants phenotype
RNA interference
Synthetic double-stranded RNA molecules matching the sequence of a particular gene are
used to break down or block the gene’s mRNA
SNPs (single nucleotide polymorphisms)
genetic markers
totipotent cell
on that can generate a complete new organism
stem cell
unspecialized cell that can reproduce itself indefinitely and differentiate into specialized cells of one or more types
embryotic stem cells can make:
- liver cells
- nerve cells
- blood cells
adult stem cells
- blood cells
gene therapy
alteration of an afflicted individuals genes (very risky)
-vectors are used for delivery of genes into specific types of cells
transgenic
made by introducing genes from one species into the genome of another animal
-these animals are pharmaceutical “factories” producers of large amounts of substances for medical uses
genetic profile
individuals DNA sequence
short tandem repeats
variations in the number of repeats of a specific DNA sequence
Ti plasmid
most commonly used vector for introducing new genes into plant cells
GMO
-putting pesticides on plants so animals dont eat them and thus can have a bigger output in product
