ning- express yo self Flashcards
__-__ _____ is a powerful technique for identifying specific ____ ___ within individual cells in tissue sections, providing insights into physiological processes and disease pathogenesis.
In situ hybridization; mRNA species
In situ hybridization is used to reveal the location of specific __ ___ ___ on chromosomes or in tissues. This is crucial for understanding the organization, regulation, and function of genes.
nucleic acid sequences
The key techniques for in situ hybridization currently in use include: in situ hybridization to mRNA with ___ and ___ ___, analysis with __ and ___ ____, whole ___ in situ hybridization; double detection of __ and ___ ___ __; and fluorescent in situ hybridization to detect ___ ___.
oligonucleotide and RNA probes (both radio-labelled and hapten-labelled); light and electron microscopes; mount; RNAs and RNA plus protein; chromosomal sequences
____ are the most-commonly used bacterial cloning vectors
Plasmids
Insertion of DNA into a bacterial plasmid: - Plasmic cloning vectors contains a site that allows DNA fragments to be inserted, for example a multiple cloning site or ____ which has several commonly-used restriction sites to which ___ ___may be ligated.
polylinker; DNA fragments
Insertion of DNA into a bacterial plasmid: After the gene of interest is inserted, the plasmids are introduced into bacteria by a process called ____
transformation
Insertion of DNA into a bacterial plasmid: These plasmids contain a selectable marker, usually an ___ ____ ___, which confer on the bacteria an ability to survive and proliferate in a selective growth medium containing the particular antibiotics
antibiotic resistance gene
Insertion of DNA into a bacterial plasmid: The cells after transformation are exposed to the selective media, and only cells containing the ___may survive
plasmid
Insertion of DNA into a bacterial plasmid: In this way, the ____ act as a filter to select only the bacteria containing the plasmid DNA. The vector may also contain other __ __ or __ __ to facilitate selection of plasmid with cloned insert. Bacteria containing the plasmid can then be grown in large amounts, harvested, and the plasmid of interest may then be isolated using various methods of plasmid preparation.
antibiotics; marker genes; reporter genes
A ____ is a small DNA molecule within a cell that is physically separated from a chromosomal DNA and can replicate independently
plasmid
__ ___ is a specific type of enzyme that facilitates the joining of DNA strands together by catalyzing the formation of a phosphodiester bond. When purified it is used in gene cloning to join DNA molecules together to form ___ ___.
DNA ligase; recombinant DNA
In molecular biology, a ____ is a collection of DNA fragments that is stored and propagated in a population of micro-organisms through the process of molecular cloning. There are different types of them, including ___ ___ (formed from reverse-transcribed RNA), __ ___ (formed from genomic DNA) and __ __ ___ (formed by de novo gene synthesis where alternative nucleotides or codons are incorporated).
library; cDNA libraries; genomic libraries; randomized mutant libraries
In genetics, ___ ___ is single-stranded DNA synthesized from a messenger RNA (mRNA) template in a reaction catalysed by the enzyme reverse transcriptase
complementary DNA (cDNA)
__ ___ is a method of investigating the sequence specificity of DNA-binding proteins in vitro. Why is it used?
DNA foot printing; This technique can be used to study protein-DNA interactions both outside and within cells
In DNA footprinting- A protein that specifically binds a region within the DNA template will __ __ __ it is bound to from the cleavage agent. Run both samples side by side on a __ __ __. The portion of DNA template without protein will be cut at random locations, and thus when it is run on a gel, will produce a __ __ __. The DNA template with the protein will result in __ __ with a break in it, the “footprint”, where the DNA has been ___ from the cleavage agent. DNA footprinting helps to determine how DNA is __ .
protect the DNA; polyacrylamide gel electrophoresis; ladder-like distribution; ladder distribution; protected; transcribed
__ __ __ is a molecular biology method that is used to make specific and intentional changes to the DNA sequence of a gene and any gene products, involving:
___ separation, __ __ primer containing desired mutated sequences; strand completion by __ ___ and __ __, and
introduction into cells followed by replication and segregation into __ __.
Site-directed mutagenesis; Strand; Synthetic oligonucleotide; DNA polymerase and DNA ligase; daughter cells (can create sequence that is changed at only 1 nucleotide
__ ___ (also, replacement strategy based on homologous recombination) is a genetic technique that uses homologous recombination to change an endogenous gene. The method can be used to: (4)
Gene targeting; delete a gene, remove exons, add a gene, and introduce point mutations
In gene targeting ___ __ __ growing in tissue culture are used to introduce a DNA fragment containing __ __ into many cells, Let each cell grow to form a ___, then test for to see if it has the DNA fragment has replaced one copy of the __ __.
Embryonic stem cells, altered gene; colony; normal gene
Inject embryonic stem cells into early embryo (Embryonic stem cells contain one copy of target gene)
transcription is __ to ___. The three phases are __, __, and ___.
DNA to mRNA; initiation, elongation and termination.
