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Genetic Engineering (MCB5426) exam 1 definitions

Terms in this set (60)

Daughter Duplex

result of replication. identical to original, has one parent strand and one newly synthesized strand (daughter)

enzyme that catalyzes DNA replication.

Alpha-Phosphate

first phosphate in the tri-phosphate NTP strand. Used in nick translation and random priming

Nick Translation

way to make labeled probes. DNase 1 will nick DNA every so often (excise NT) and then DNA polymerase will bind to nick and synthesize new strand from nick onward. 'Hot' base alpha-labeled gets added into new strand

non coding sequence of RNA transcript (or DNA encoding it) that will get spliced out during RNA processing

Directional Cloning

inserting a target gene into vector in a specific and known direction. ligation of an insert into vector in known orientation using 2 different restriction enzymes that yield 2 different sticky ends

coding sequence of DNA (and corresponding RNA) that is a part of the final mature mRNA sequence

Shotgun Cloning

method used for sequencing long DNA strands. DNA is first shredded into smaller fragments which can be sequenced individually. used for human genome project

Alternative Transcript/Splicing

allows production of alternate proteins from the same set of introns and exons, tissue or time dependent.

small interfering RNAs, defense against genomic invaders aka viruses

complimentary DNA - DNA that is made from an RNA template

DNA reading frames

a way of diving a sequence of nucleotides in a nucleic acid molecule into a set of consecutive triplets (codons) that would be translated into amino acids. 6 for any given strand of DNA

sticky ends on DNA. there are extra nucleotides on one side of the ds DNA making it partially ss DNA. Stretch of unpaired NTs

Electroporation

utilization of a high voltage electric current that is passed through a bacterial suspension for several milliseconds. 'zaps' cells to open up membrane and insert vectors

Gel Electrophoresis

way to visualize DNA by size. uses charge and a salt buffer to separate DNA by size as it moves through an agarose or acrylamide gel

Calf intestine phosphate

CIP - catalyzes removal of phosphate groups from 5' ends of DNA stands and adds OH group. Removes y-phosphate

Primary transcript

single stranded RNA molecule that is synthesized by DNA transcription. later modified for translation. contains introns, exons, poly A tail, G cap

5' end of sequence that pairs with sequence near 3' end of small rRNA subunit in prokaryotic translation

ability to bind DNA and protect it from external nucleases

when bacterial genes for a particular metabolic pathway are contiguous. contains a promoter, large mRNA, close placement of start and stop codons, and 2 DNA binding sites specific for RNA polymerase

when phages enter the bacterial cell and produce even more phage which ends up lysing and killing the cell - virulent

TATA box, allows RNA polymerase to recognize DNA

Lysogenic Cycle

when a phage enters a cell and integrates into the chromosome to become quiescent

Initiation Complex

eukaryotic translation. forms by assembly of ribosomal subunits and initiator tRNA at the start codon of mRNA

follows formation of transcription complex. process by which amino acids are added onto one another in a chain based on mRNA sequence being read by ribosome. forms protein

lactose operon of E.coli. consists of 3 structural genes, promoter, terminator, regulator, and operator. encodes for ß-gal which is an intracellular enzyme that turns lactose into glucose and galactose

small molecule that will bind to repressor or activator to help regulate gene transcription. can be a corepressor, activator, or repressor

enzyme responsible for fixing the sugar backbone of DNA. fixes singe stranded nicks in DNA chain, requires ATP. reforms phosphodiester bond in backbone of adjacent nucleotides

small molecules that are often bound to DNA to prevent transcription. when effector binds to repressor, allows transcription to occur. can also be inactive in resting state but become active in the presence of effector to stop transcription

introduction of DNA into a bacterial cell host. Genetic alteration of a cell resulting from uptake of exogenous genetic material. when foreign DNA is taken up into a cell

uptake of entire functional plasmid into functional cell

non-chromosomal material that is double stranded, circular, and naturally occurring. Replicons that are inherited in extrachromosomal state. intact, CCCDNA

technique that measures the degree of genetic similarity between pools of DNA sequences, can determine genetic distance between organisms. attaching ss DNA strands together with H bonds to find gene of interest

bacterial artificial chromosome, plasmid based vector that is maintained as a bacterial chromosome. can hold 50-300 kB inserts. maintained in low copy #

human artificial chromosome. therapeutic vector that has a large DNA carrying capacity. ranges from .7-400mB. microchromosome that can act as a new chromosome in a population of cells. uses same essential elements as normal chromosome (replication origin, centromere, telomere). entirely separate chromosomes, so no insertional mutants arise

labeling method where DNA polymerase is modified not to have exonuclease activity. denature, add oligos, and allow hybridization to proceed, then start DNA polymerization with Klenow and a hot nucleotide. labels small daughter strands of DNA sequence that are 200-500 bp in length

best example of a temperate bacteriophage. has cos ends, you can replace the middle with your DNA of choice

covalently closed circle DNA. intact plasmids

transfer of DNA to a membrane matrix which is then used in hybridization to find gene of interest

Multiple Cloning site

a short sequence of DNA that contains many restriction sites. standard feature of engineered plasmids. single site for a large number of restriction endonucleases, preferably in genes with a readily scorable phenotype

Transfer of RNA to a membrane matrix. RNA gel, blot as for southern. single stranded sequences.

helps package DNA into bacteriophage head. at each cos site, DNA will be cleaved so you only have one copy of genome per head. creates sticky ends

transfer of protein bands to a membrane matrix, use antibodies to detect antigens

plasmid sequences joined to cos sites of lambda phage, small.

when different enzymes recognize the same DNA sequence. may cut at different sites

a complimentary primer to a poly-A tail providing a free -OH end that can be extended by reverse transcriptase. Deoxy T residues

Dideoxynucleotide

modified nucleotide that lacks the 3' OH group. will stop synthesis

Polymerase used in PCR reactions. Comes from bacteria found in hot springs

end of a DNA double helix at which a few unpaired NT of one strand extend beyond another

two phosphate groups linked by esterification. released in synthetic steps during protein and nucleic acid elongation.

Origin of replication. DNA sequence that signals replication to begin

library that is dependent on knowing the size of the restriction fragment on which a particular gene is located

microRNAs. regulate endogenous genes. removes stem-loop precursors with imperfect double strandedness. post transcriptional regulation of genes

Recognition Sequence

4-6bp stretch of DNA that a restriction enzyme will bind to and cut

how different genes interact with each other to produce a desired phenotype

genetic engineering

any alteration of an organisms genes for practical purposes

functional genomics

makes use of data gathered by genome sequencing projects to describe genome function. uses high throughput techniques to describe function and interactions of genome

directed evolution

repetitive cycles of DNA manipulations on a defined gene or defined set of homologous genes resulting in gene mutations, followed by screening for desired phenotypes

specific nucleotide sequence that is transcribed into RNA. does not have to be translated

when a phage can do both lytic and lysogenic cycles

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