Ch. 20 APBIO ( DNA Technology and Genomics) Flashcards

DNA
Terms Definitions
Gene Therapy
The insertion of working copies of a gene into the cells of a person with a genetic disorder in an attempt to correct the disorder, for it to be permanent it must involve the cells that will proliferate throughout a persons lifetime(like BONE Marrow Cells)
Human Genome Project
An international effort to map the complete human genetic code. This effort was essentially completed in 2001, though analysis is ongoing.
In Vitro Fertilization
The most common assisted reproduction procedure, in which a woman's eggs are mixed with sperm in culture dishes (in vitro) and then carefully inserted into a woman's uterus.
Golden Rice
Vitamin A deficiency is a serious health issue, so 2 genes from daffodils and 1 from bacteria were inserted, with added genes rice plants synthesize betacarotene, whtn the rice is eaten it is converted to Vitamin A
Recombinant DNA
Genetically engineered DNA made by recombining fragments of DNA from different organisms
Genetic Engineering
The direct manipulation of genes for practical purposes, which include the manufacture of protein products(like Hormones and blood clotting factors), by using this technological approach you can make recombinant DNA and then reintroduce it into cultured cells
Biotechnology
A form of technology that uses living organisms, usually genes, to modify products, to make or modify plants and animals, or to develop other microorganisms for specific purposes.
Metazoa
Multicellular animals having cells differentiated into tissues and organs and usually a digestive cavity and nervous system
DNA Microarray
Technique used to screen a single sample for a vast range of different nucleotide sequences stimultaneously; it is often used to study gene expression
Gene Cloning
The process of isolating a gene sequence in the genome of an organism and inserting the gene sequence into a plasmid vector for production in large numbers.
Pedigree
a diagram that shows the occurrence of a genetic trait in several generations of a family
Bacterial Plasmid
A Circular DNA molecule found in bacteria which can be inserted with foreign DNA.
Used to mass produce insulin and human growth hormone.
Recombinant Bacterium
Insert foreign DNA into a plasmid and then put the plasmid in a bacteria so it has foreign DNA too
Clone
a group of genetically identical cells or organisms derived from a single cell or individual by some kind of asexual reproduction
Cloned Genes
Are useful for two main purposes, they can be used to make many copies of a particular gene and can produce a protein product, which may endow an organism with a new metabolic function such as pest resistance.
Restriction Enzymes
Also called restriction endonucleases, were first discovered in the late 1960' s and have made genetic engineering possible. They are found naturally in Bacteria and protect against foreign organisms. Is very specific and only recognizes a particular DNA sequence
Restriction Fragments
DNA segment resulting from cutting of DNA by a restriction enzyme at a restriction site (Usually Many
Sticky Ends
Short, single-stranded regions of DNA that came from broken double-stranded palindromic DNA.
Restriction Sites
The DNA sequence that is recognized by a restriction enzyme; the restriction enzyme cuts at this sequence, generating two DNA fragments (Usually 4 to 8 Nucleotides) , and because it is so small, restriction fragments are cut out at many places.
DNA Ligase
An enzyme that catalyzes the formation of covalent bonds that close up the sugar phosphate backbone after the sticky ends form complementary hydrogen bonds(because these are only temporary)
Cloning Vector
An agent used to transfer DNA in genetic engineering. A plasmid from bacteria that moves recombinant DNA from a test tube back into a cell is an example , or it can be a virus that transfers recombinant DNA by infection.
ampR
Antibiotic resistance gene, found in vectors derived from natural episomes (plasmids) and bacterial genome.
Ampicillin
Mech same as penicillin. Need a B-lactamase inhibitor. Used against Gram+ and HELPS (Haemophilus Influenza, E. Coli, Listeria, Proteus mirabilis, and Salmonella). SE: Pseudomembraneous Collitis (because of removal of gut bacteria). IV Form.
LacZ
Gene part of the lac operon. If gene is intact, produces a product that can break down lactose and Xgal, and the colony WILL be blue. If not intact, it will NOTbreak down Xgal, colony will be white.
X-Gal
A Chemical similar to lactose that turns dark blue when cleaved by beta-galactosidase
Agar
a gel-like polysaccharide compound used for culturing microbes; extracted from certain red algae
10^5
The amount of cells that need to be present sothat they can be seen on an agar plate.
Nucleic Acid Hybridization
a form of DNA technology used to detect specific DNA or RNA sequences based on their ability to anneal to nucleic acid probes , if it know part of the neuclotide sequence of the gene of interest., then we could synthesize a probe that is complementary to it, which will then be labeled after it hydrgen bonds so that we could track it. KEY to this process is denaturazation of the Cell's DNA.
Nucleic Acid Probe
In DNA technology, a labeled single-stranded nucleic acid molecule used to locate a specific nucleotide sequence in a nucleic acid sample. Molecules of the probe hydrogen-bond to the complementary sequence wherever it occurs; radioactive or other labeling of the probe allows its location to be detected.
Genomic Library
A set of thousands of DNA segments from a genome, each carried by a plasmid, phage, or other cloning vector
Phage Library
Most efficient genome library, 160,000 are needed to make up human genome, made by the backaging of the recombinant DNA into bacterial cells, where they replicate and produce new phage particles, which are stored as a selection of phage clones.
Plasmid Library
Cannot contain much info, 700,000 are needed to make the human genome, made by a collection of bacterial cells each of which containing copies of a particular genome fragment
Complementary DNA
A DNA molecule made in vitro using (mRNA) as a TEMPLATE and the enzyme reverse transcriptase . Also called (cDNA) molecules , they corresponds to a gene, but LACK the introns present in the DNA of the genome. Useful for studying the specialized function of a particular cell type.
Reverse Transcriptase
An enzyme encoded by some certain viruses (retroviruses) that uses RNA as a template for DNA synthesis.
Expression Vector
A cloning vector that contains the requisite bacterial promoter just upstream of a restriction site where a eukaryotic gene can be inserted, allowing the gene to be expressed in a bacterial cell. Allowing for the synthesis of many eukaryotic proteins in bacterial cells.
YACs
yeast artificial chromosomes. They are linear, like normal yeast chromosomes, not circular like plasmids. They can be used to clone very large pieces of DNA, up to about 500 kbp. They are not used much anymore, as BAC clones have technical advantages., Can surmount the differential transciption between prokaryotes and Eukaryotes.
Electroporation
A technique to introduce recombinant DNA into cells by applying a brief electrical pulse to a solution containing cells. The electricity creates temporary holes in the cells' plasma membranes, through which DNA can enter.
Agrobacterium
A bacterium that forms Galls in plants and transfers some of its genes into plant chromosomes through conjugation
Polymerase Chain Reaction
A method of producing thousands of copies of DNA segment using the enzyme DNA polymerase(Usually From Archaea), it is especially useful when the amount of DNA present is very scant or impure, because it is a quicker(a couple billion in a couple of hours) and more selective. Requires Double Stranded DNA containing target sequence to be copied, heat resistant DNA Polymerase, all 4 Nucleotides, and two short,single stranded DNA molecules which will serve as primers. Problem becose of occasional errors.
Restriction Fragment Analysis
DNA fragments produced by restriction enzyme digestion of a DNA molecule are sorted by gel electrophoresis; is useful for comparing two different DNA molecules such as two allels for a gene; used to prepare pure samples of individual fragments
Gel Electrophoresis
A procedure used to separate and analyze DNA fragments by placing a mixture of DNA fragments at one end of a porous gel and applying an electrical voltage to the gel, the ones that are largest( or least negative) will move the slowest and farthest , while the smallest will do the exact opposite. These bands are NOT visible until a marker is added.
Southern Blotting
A technique that enables specific nucleotide sequences to be detected in a sample of DNA. It involves gel electrophoresis of DNA molecules and their transfer to a membrane (blotting), followed by nucleic acid hybridization with a labeled probe. (Must use an alakine solutiojn, Gel, Nitrocellulose, and a heavy weight to pull the solution through the cell. Most useful application is its use to identify the heterozygote carriers of mutant alleles associated with genetic diseases.
RFLPs
Differences in homologous DNA sequences that are reflected in different lengths of restriction fragments produced when the DNA is cut up with restriction enzymes
Physical Mapping
Assign genes to a particular locations using measurements that are a true reflection of the physical distance between the genes, usually by the number of base pairs along the DNA, which are then arranged so that they overlap
Linkage Mapping
Genes on a chromosome are arranged in a linear array, and the physical distance between them dictates the frequency of crossing over between them. The greater the physical distance,
the greater the frequency of crossing over. Made after a cytogenic maps are constructed by in situ hybridization.
Cytogenetic Map
a map of a chromosome that includes the positions of genes relative to visible chromosomal features, such as stained bands
BAC
Also called bacterial artificial chromosomes, which can clone much larger pieces of DNA but have lower copy number
Fredrick Sanger
the pioneer in determining the amino acid sequence of proteins, through his use of Dideoxy Chain Termination Method for Sequencing DNA .
Dideoxy Chain Termination Method
A method of determining a sequence of nucleotides in any CLONED DNA fragments up to 800 base pairs in length which can be determined rapidly by using a nested set of DNA strands complementary to the original DNA fragment, each that starts with the same primer and ends with Dideoxyriboneuclotide.(ddNTP), which terminates a growing DNA strand because it lacks a (OH-), and because they are tagged with a flourescent label , it determines the ending of the sequence which can be then used to sequence the entire DNA. (Developed by Fredrick Snger)
Hemophilus Influenzae
Gram- bacillus responsible for 5% of all bacterial meningitis cases,l also first complete genome sequenced by Vente and Celera Genomics
Celera Genomics
privately owned biotech company (Craig Venter)
techniques: relied on newer/faster DNA sequences & programs
*Hare
Craig Venter
entreprenuer who worked for Celera. Developed the "Shotgun" sequencing method, wanted people to pay to view genes in database, raced against Francis Collins to Finish Genome
Shotgun Approach
• As genomes get larger, piecing together information gets more difficult.
• Typically used for smaller genomes, like bacteria.
• Early step is to fractionate DNA to that the fragments of the particular size are used.
o This is done by using a sonificator which breaks
the DNA into particular sizes that can be shown
on a gel and extracted and purified. The
end-sequences of DNA inserts are obtained and
put into computer to sequence them.
• Organizing the clones, you generate probes from the clones creating end-sequences and apply it to the entire genome. If the probe sticks to numerous places then you can attempt to say that they line up to each other in the genome.
Genomics
study and comparison of genomes within a single species or among different species
Expressed Sequence Tags
certain short sequences that correspond to sequences present in known mRNA, are catalouged in a computer, which identifies sequences that are new product coding genes.
In Vitro Mutagenesis
A technique to discover the function of a gene by introducing specific changes into the sequence of a cloned gene, reinserting the mutated gene into a cell, and studying the phenotype of the mutant. RNAi is more effective and faster.
RNAi
RNA interference; injecting double stranded RNA into a cell turns off expression of a gene with the same sequence as the RNA, useful in assesing differential expression of genes.
DNA microarray assays
A method to detect and measure the expression of thousands of genes at one time. Tiny amounts of a large number of single-stranded DNA fragments representing different genes are fixed to a glass slide. These fragments, ideally representing all the genes of an organism, are tested for hybridization with various samples of cDNA molecules.
Proteomes
the full protein sets encoded by genomes
Proteomics
the study of all of an organism's proteins, including its identity, structure, interaction, and abundance
SNPs
a DNA sequence variation occurring when a single nucleotide — A, T, C, or G — in the genome (or other shared sequence) differs between members of a species (or between paired chromosomes in an individual). For example, two sequenced DNA fragments from different individuals, AAGCCTA to AAGCTTA, contain a difference in a single nucleotide. In this case we say that there are two alleles : C and T. Frequency may vary with ethnicity
RT-PCR
is a variant of polymerase chain reaction (PCR), a laboratory technique commonly used in molecular biology to generate many copies of a DNA sequence, a process termed "amplification". An RNA strand is first reverse transcribed into its DNA complement (complementary DNA, or cDNA) using the enzyme reverse transcriptase, and the resulting cDNA
SCID
Severe combined immuno-deficiency. In this disorder both B-cells and T-cells are absent and therefore such babies are highly susceptible even to minor infections.
HGH
(Human growth hormone) also known as somatotrophic hormone is responsible for the growth of long bones, muscles and viscera.
Tissue Plasminogen Activator
converts PLASMINOGEN to PLASMIN in the presence of fibrin ,may lose fibrin specificity at high doses which disolves clots(Like in the case of a heart attck) and bleeding is common
DNA Fingerprints
Compares repeated sections of genes that have little to no known function, but vaary widley from one person to another(STRs). gel electrophoresis used to separate fragments, the specific repeats are then labeled using radioactive probes producing bands to e compared
STRs
Short Tandem Repeats, regions of a DNA molecule that contain short segments consisting of three to seven repeating base pairs
Transgenic
term used to refer to an organism that contains genes from other organisms
Ti Plasmid
a plasmid of a tumor-inducing bacterium that integrates a segment of its DNA into a chromosome of a host plants. frequently used as a vector for genetic engineering in plants. Comes form Agrobacterium tumefaciens
Genetically Modified Organisms
organisms whose genetic code has been altered by artificial means such as interspecies gene transfer
Transgene
foreign gene that is transferred into target cell or tissue
T DNA
a strech of DNA found in a Ti plasmid, INJECTED IN THE CHROMOSOMAL dna of its host.
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