In Vivo Cloning

Flashcards about In Vivo Cloning Lecture Notes

Genetic engineering is __

The modification and manipulation of an organism's genes using technology.

__ involves recombinant DNA.

DNA that is a combination of originally different DNA molecules put together in the lab, usually from different biological sources, that are not found together in nature.

Molecular cloning is __

Making many identical copies of a DNA sequence.

In vitro means __

In glass…the study takes place in a test tube, plate, or flask

In vivo means __

In life…the study takes place in a living organism

Restriction enzymes __

They are used in the lab to cut DNA into pieces.

Digestion is __

Using a restriction enzyme to cleave DNA.

Restriction enzymes break covalent bonds in the sugar-phosphate backbone. Specifically, the break is __

The break between the 3′ oxygen of one nucleotide and the 5′ phosphate of the next one.

A palindrome is__

A word, phrase, or sequence that reads the same backward as forward, like madam.

Sticky ends __

Generated by cuts with the same RE (restriction enzyme) have complementary base pairs and will naturally bind to each other.

Recombinant DNA involves stitching DNA fragments into __

DNA fragments are generated using restriction enzymes or PCR. DNA fragments and vectors are sealed together by the enzyme DNA Ligase.

A plasmid is__

An extrachromosomal double-stranded DNA molecule that replicates autonomously in bacterial cells.

__ is a selectable marker

Ampicillin resistance is a type of selectable marker that allows us to identify bacteria that have successfully taken up the plasmid.

The in vivo method of cloning involves introducing __ into bacteria

A recombinant DNA molecule is transferred to a host bacterial cell to produce many copies.

Transformation is __

A process by which bacteria take up plasmids.

CaCl2 is used to chemically prepare __

Neutralizes the negative charges on the cell surface and DNA, reducing electrostatic repulsion; Also destabilizes the bacterial cell membrane, creating transient pores that facilitate DNA entry.

Heat shock is __

Involves exposing bacterial cells to a brief period of high temperature (usually 42°C), followed by a return to a cooler temperature (usually on ice).

Transformants are __

Those bacteria that have successfully taken up the plasmid.

Transformants are identified by __

Growing transformants on a plate containing the antibiotic that the plasmid encodes resistance to.

Recombinant DNA involves stitching DNA fragments into __

DNA fragments are generated using restriction enzymes or PCR. DNA fragments and vectors are sealed together by the enzyme DNA Ligase.

The goal of a diagnostic digest is __

To cut a recombinant plasmid into specific-sized pieces and analyze the resulting fragments by gel electrophoresis.

A positive control in a restriction digest __

Confirms that the reaction is working as expected and that the enzyme is active.

Agarose gel electrophoresis __

Separates DNA products by size and charge.

Agarose gels __

Act as sieves that separate DNA fragments by size.

Agarose powder is __

A polysaccharide gelatin that is isolated and purified from agar-bearing seaweed.

TAE buffer__

Tris-acetate-EDTA is a running buffer commonly used to separate nucleic acids.

Fluorescent DNA stains __

Bind to the DNA double helix.

Glycerol __

Weighs down samples.

DNA ladders __

Are used to measure sizes of DNA molecules

Add just enough TAE buffer to the chamber to cover the gel because __

Buffer provides ions for conducting current during electrophoresis

Negatively charged DNA samples __

“run to red”

A band on a DNA gel is __

each band on the gel contains DNA molecules that are a specific size