DNA in the lab Flashcards
What does the melting temperature of DNA depend on?
The AT:CG ratio
A and T have 2 H bonds, C and G have 3 H bonds
What structures are present in the double helix structure as a result of the asymmetrical helices?
Major and minor grooves
What causes the major and minor grooves?
The orientation of bases as the two glycosidic bonds that a base pair can form are not directly opposite each other
What does it mean that RNA viruses break the central dogma?
They have genomes encoded in RNA
How do retroviruses work?
Retroviruses such as HIV use reverse transcriptase to convert RNA genomes to DNA then use host DNA replication and transcription systems systems using an RNA-dependent DNA polymerase
How do RNA viruses work?
RNA viruses such as Ebola and COVID use special polymerase enzymes that allow them to make RNA from their RNA genomes using an RNA-dependent RNA polymerase
What are the two types of information carried by DNA?
Coding information (sequence dependent) - by complementary base base-pairing, requires local dissociation of double helix for transcription/replication Protein-DNA binding information (sequence dependent but dependent on features in major/minor groove)
Where do polymerases add new nucleotides and so what direction do strands form in?
They add to the 3’ position
All strands form in 5’ to 3’ direction
What determines start site and direction of an RNA polymerase?
A promoter eg. TATA
What are enhancers? (also called regulatory elements REs)
Regulatory DNA sequences that can enhance gene transcription
They are non-directional, variable distances from the promoter and may be tissue-specific
What are typical modern cloning vectors?
Plasmid vectors or phage vectors
What is a plasmid vector?
Has a polylinker site made up of restriction enzyme sites which allows plasmid to be cut open and have region inserted
All other restriction enzyme sites are removed
Polylinker sometimes inserted into LacZ
Antibiotic resistance gene allows selection to isolate only bacteria that have taken up plasmid
What is a phage vector?
Useful for larger DNA
Can be cut open with EcoR1
Constructs inserted
Used for infecting bacteria
