Nucleic Acid Structure Flashcards
Posttranslational Histone Modifications
+Reversible
+Influence chromatin structure and function
+Govern gene activity (transcription)
Acetylation
Lysine E-amino group
Methylation
Lysine E-amino group
O-Phosphorylation
Serine, Threonine
N-Phosphorylation
Histidine, Lysine
Nucleosome
A structural unit of a eukaryotic chromosome, consisting of a length of DNA coiled around a core of histones
How many times does DNA wrap around a nucleosome?
Two times
Which histone is not a part of nucleosome core?
H1
Histone octamer
The 8 protein complex found at the center of a nucleosome core particle.
Which histones make up the histone octamer complex?
+Two H3 and H4
+Two H2A-H2B dimers
What modulates transcription?
DNA packing
During interphase what is the percent of DNA in heterochromatin?
10% (transcriptionally inactive)
During interphase what is the percent of DNA in euchromatin?
90% (10% in open state, 90% in intermediate packed state)
How is DNA degraded?
By breaking phosphodiester bonds
Enzymes for degradation
DNases
RNases
Exonucleases
Endonucleases
Non-enzymatic degradation
Very high heat (>95C)
How is DNA denatured?
By breaking H bonds
Enzymes for denaturing
Helicases
Non-enzymatic denaturation
High heat (
Hyperchromicity
The increase of absorbance of a material; occurs when the DNA duplex is denatured; ssDNA absorbs ~40% more than dsDNA
Hypochromicity
A decrease of absorbance of a material; dsDNA absorbs ~40% less than free nucleotides
DNAs maximal absorbency
A260
Melting temperature midpoint (Tm)
Temperature at midpoint of denaturation reaction
Factors affecting Tm
+ G+C content
+ Ionic strength
+ Presence of chaotropic agents
SSC Buffer
Standard Sodium Citrate
Annealing
Forming H-bonds between two ssDNA to yield dsDNA ; depends of complementarity between ssDNA1 and ssDNA2
Renaturation
Annealing of ssDNA from one source to reform dsDNA
Hybridization
Annealing of ssDNA from 2 sources to form hybrid dsDNA
Similarity
Proportion of bases identical in sequence
Complementarity
Portion of bases in 2 ssDNA complementary to each other
Solution hybridization method 1
Determine A260 (hyperchromicity)
Solution hybridization method 2
Hydroxyapatite chromatography
ssDNA elutes with ___ Pi buffer
low
dsDNA elutes with ___ Pi buffer
high
Filter hybridization
Establishes whether a solution of denatured DNA (or RNA) contains sequences complementary to the strands immobilized on the filter
Factors affecting reassociation kinetics
+ Temperature (Tm=25)
+ Monovalent cations block reassociation
+ High temp weakens secondary structure
+ Time and concentration to ensure collisions
+ Size and length of DNA chain (usually shear DNA to 300 bp)
Eukaryotes exhibit 3 components (in reassociation kinetics)
+ Fast
+ Intermediate
+ Slow
Slow component repetition
Single copy
Fast component repetition
Many copies (500,00 freq)
Intermediate component repetition
Some copies (350 freq)
C0t1/2 =
Fraction of genome X C0
Complexity of unknown =
C0t1/2 of unknown X Complexity of standard /// C0t1/2 of standard
Chemical complexity (genome size) =
Nucleic acid /// Nucleus
Stringency
Control amount of mismatch permitted in reaction
F (repetition frequency) =
Chemical complexity /// kinetic complexity
