Notes 1 Flashcards
Purines
Two fused rings
DNA
Has a T
2’ carbon only has H
Bases named as nucleosides
Adenosine Guanosine Cytidine Thymidine Uridine
Bases named as Nucleotides
Adenylic acid Guanylic acid Cytidylic acid Thymidylic acid Uridylic acid
Types of bond between bases in a double helix
Noncovalent (H-bond)
Melting temp of DNA
75 Degrees
What causes spaces betwen the bases where we could insert something?
Hydrophobic interaction
Interchalation
Ethidium bromide can slip into DNA molecule between bases.
Major groove of DNA
22 Angstroms
Minor grove of DNA
12 Angstroms
What defines the region of a gene
Length of RNA
What defines the coding region?
Protein
Explain the process of splicing
Pre-mRNA contains an intron. The intron is cut out. It travels through a nuclear pore into the cytoplasm to be translated by a ribosome.
Trans acting
Can act on any copy of the gene in the cell
Cis acting
sequences act on same sequence/chromsome.
What are trans acting?
Proteins
What are cis acting?
sites on DNA
Nucleases
hydrolyze an ester bond within a phophodiester bond
Phosphatases
Hydrolyze the ester bond in a phosphomonester bond
Restriction endonucleases
Recognize specific short sequences of DNA and cleave duplex near or in the recognition site.
Where does Eco RI cut?
between the G and the A
Palindrome
spelled the same backwards and forward
Endonuclease
nucleases that cleave phosphoester bonds within a nucleic acid chain. Cuts INSIDE strand.
Exonuclease
cleave phosphoester bonds one at a time from the end of a chain.
Migrate in and cut from OUTSIDE
Vector
Required for cloning to move DNA
Recombinant DNA
a DNA molecule that has been created by joining together two or more molecules from different sources
Subclone
process of breaking a cloned fragment into smaller fragments for further cloning
MCS (multiple cloning site)
Region of DNA in a vector with lots of places to insert foreign DNA (multiple restriction endonuclease sites)
Selectable markers in cloning
LacZ
Ampr
Ori
Amp r
Ampicillin resistance
Ori
Origin of rep in cell residing in
lacZ
betagalactactasidase
Colonies turn blue with the right substrate
Transformation
the acquisition of new genetic material by incorporation of added exogenous, nonviral DNA
Substances that contribute to blue color of plate
IPTG and X gal
Bacterial artifical chromosome (BAC)
300 kB
F plasmid
Yeast artificial chromosome (YAC)
3000 kB
origin, centromere, telomere
Can be propagated in more than one type of host cell
Shuttle vector
Expression vector
contain promoters that allow transcription of any cloned gene
Reporter genes
Used to measure promoter activity or tissue specific expression
Good way to report on functionality
4 methods to introduce DNA into target cell
- Viral vector by infection
- Liposomes that fuse with the membrane
- Microinjection introduces DNA directly into the cytoplasm or nucleus
- Nanospheres can be shot into the cell by a gene gun
Gel electrophoresis
separates DNA fragments by size, using an electric current to cause the DNA to migrate toward a positive charger
Larger fragments take longer to get through gel.
Linear pieces take longer, supercoiled can speed through.
Plamsid
10 kB limit
Phage
20 kB limit
Cosmid
48 kB limit
