Gene regulation, Mutations and Biotechnology Quiz Review

Restriction endonucleases

naturally found in bacteria
defend themselves against viruses
destroys viral DNA by cutting it apart
cuts ALWAYS occur at specific sequences of DNA that are NOT FOUND in the bacteria cell’s own DNA

where do restriction endonucleases normally cut?

• palindromic sequences (sequences where the DNA is read the same forwards and backwards)

• results in sticky ends

what are sticky ends?

• short, single-stranded unpaired nucleotides

• they have different lengths

• they are “sticky” because they can pair with a completely different overhang on another fragment

plasmids

• small, circular, double stranded DNA molecule

• “extra” DNA that is distinct from the bacterial cell’s chromosome

• often carry genetic advantages (antibiotic resistance)

how can restriction enzymes be used as scissors?

• cut DNA and insert new genes (genetic engineering, GMO’s)

• cut DNA and separate it by size (DNA fingerprinting)

how can plasmids be used as gene delivery vessels?

• to insert genes into bacterial cells so that they can make proteins

• bacteria can make human insulin for diabetes patients

• Multiple genes can be inserted. Often, this includes a gene that codes for a protein that glows! This is useful because any bacterial colonies that glow, must have  successfully taken up the plasmid. It lets scientists ‘see’ that DNA was inserted effectively.

Gel electrophoresis

• separating DNA by size

• DNA has a negative charge

• a DNA sample is placed at the negative side of the gel

• when the machine is turned on, the DNA is attracted to the positive side, but the gel’s pores slow it down

• the bigger the DNA, the slower it will move

• so, in a given time frame, big pieces of DNA don’t move as far across the gel

polymerase chain reaction

• Denaturation: Heat to 95 degrees to separate the DNA strands.

• Annealing: Cool to -68 degrees for primers to bind to the target region.

• Extension: Heat to 72 degrees for Taq Polymerase to build the new DNA strand.