1 Genetic Material

• DNA → RNA → Protein

• 1. Adenine
• 2. Guanine

(pure as gold; are double ringed, relatively large)

• Nucleotides: base, sugar, phosphate(s)

• Nucleosides: base & sugar ONLY

1. purine or pyrimidine bases linked via N-glycosidic bond to

2. a 5 carbon cyclic sugar

3. a phosphate esterified to the hydroxyl on carbon 5 of the sugar

(Nucleotides also occur in activated diphosphate and triphosphate forms)

a covalent bond between a sugar & N group of a purine or pyrimidine base

(what was an NH₂ turns into an —N— )

JUST the bases linked to a 5 carbon cyclic sugar

(S comes before T, so you can remember it that way, that nucleoSides contain fewer components)

• polymer of deoxyribonucleotides connected via 3’ to 5’ phosphodiester linkages

• pentose-phosphate backbone

• • small groups (~5-50) of nucleotides linked via phosphodiester bonds
• • can also be called primers

• — O — H --- N —
• • holds DNA’s double helix together

• double helix is held together via H-bonds between BASES of opposite strands

• it’s specific; A binds only to T; G binds only to C

20Ä

3.4Ä

every 10 residues

in the MAJOR groove

• the larger size of the major groove makes it more accessible for interactions with proteins that recognize specific DNA sequences

because they’re lined by potential hydrogen bond donor & acceptor atoms

because the N-glycosidic bonds of a base pair are not diametrically opposite each other

• • DNA Kinase adds phosphate groups to DNA
• DNA-OH → DNA-OPO₃

• • Phosphatase removes phosphate groups from DNA
• DNA-OPO₃ → DNA-OH

• • when the DNA is twisted in the same direction as the winding of the double helix
• • “wound more tightly” (twisted in the direction of the coiling)

• • when DNA is twisted about its axis in a direction opposite to the intrinsic turns of a right-handed double helix
• • “unwound”

one that lacks supercoiling

supercoiled ones

this excess energy is available to do work (eg. separate strands of DNA)

• enzymes that catalyze the 3 step process that changes the supercoiling of DNA

• important because they can change the TOPOLOGY of DNA

1. strand cleavage (1 or both DNA strands)

2. strand passage (passage of strand through the break)

3. ligation (resealing of the DNA break(s))

during step 2 of the topoisomerase process, Topo II cleaves just one strand

during step 2 of the topoisomerase process, Topo II cleaves both strands

Camptothecin inhibits Topo I

m-AMSA inhibits Topo II [so does Doxorubicin]

the usual right handed DNA helix

• left handed helix

• short DNA molecules composed of alternating purine-pyrimidine nucleotides (especially G's & C's) adopt an alternative left handed helix

in response to negative super-coiling

• formation of Z-DNA reduces the number of negative supercoils

1. a sequence of adenine repeats

2. protein-induced bending (eg. around histones)

triplex DNA structures (H-DNA)

• can be formed by polypurine/polypyrimidine tracts (biological consequences of H-DNA are unknown)

1. sequence composition: double strands rich in G ≡ C are more stable than those rich in A = T → have higher T_m values

2. ionic conditions (NaOH can also be used to denature DNA)

they REANNEAL

eg. to hybridize a probe, lower the temperature & complimentary base pairing will occur

ssDNA (or RNA) that bind to a nucleic acid of interest via complementary base pairing

• • are derived from different sources (eg. cDNA, genomic DNA, oligonucleotides)
• are usually labeled (eg. radioactive, fluorescent)

~3 billion bp

• • average mRNA is ~ 1000 nt
• • the human genome is large enough to encode ~3 million proteins

~ 25,000

a considerable part of our genome is NOT encoding protein

Gene cluster (eg. beta globin gene cluster)

looks like a gene but isn’t functional

a piece of DNA homologous to a coding segment that does not code for a protein

sequences of DNA that are formed when mRNA is reverse transcribed into DNA & inserted back into the genome

might see poly T & complementary poly A sequences in processed pseudo-genes because a hallmark of mRNA is the 3’ polyA tail

INTRONS

1. mRNA is reverse transcribed into cDNA

2. RNase (ribonuclease) degrades the mRNA copy that was used as a (reverse) transcript

3. DNA polymerase transcribes the cDNA & creates a complementary strand

4. Integrase incorporates the dsDNA into a chromosome

DNA copies of retroviruses inserted into the chromosome

human genome contains ~1,000 proviruses; thought that 8% of the human genome is derived from retroviruses

1. transposable elements [SINEs & LINEs]

2. simple sequence repeats [SSRs]

3. satellite DNA

• a piece of DNA that can insert copies of itself in a new location within the genome

• divided into 2 main classes based on their mechanism of transposition: reverse transcription, or transposase encoded within themselves

an enzyme that allows movement of a transposable DNA element; some elements encode their own transposase

1. SINE elements

2. LINE elements

3. proviruses

4. processed pseudo-genes

~40%

among the MOST abundant sequences in the mammalian genome

eg. Alu repeats

• each ~280 nucleotides long

• 1 Alu repeat every ~5,000 bp (5 kb)

~10%

• constitute ~20% of human genome

• eg. L1 Family

• complete LINE elements are 6-8 kb long but majority are truncated

LINEs are at least at least 500 nts long

• also full-length LINE elements encode a reverse-transcriptase; it’s thought that SINE elements used LINEs’ reverse transcriptase to get into genome

NO

the presence in a population of two or more allelic variants

differences in the human genome that aren’t mutation causing

• 1. Microsatellites (very short repeats)
• 2. Minisatellites (14-500 base pair repeats)

• • 2-5 bp repeats
• • mean array size: ~100
• • present at many locations (eg. ~100,000 copies of the d(CA) repeat)
• • highly polymorphic
• • present in everybody at the same specific chromosomal locations
• • eg. GTGTGTGTGT

• • repeat lengths of 14 to 500 bp
• • mean array size is ~10 to 100
• • also highly polymorphic

• SSRs tend to be highly polymorphic
• the number of copies varies from one individual to another, allowing them to be useful biomarkers

SINEs: 10% (Alu specifically)

LINEs: 20%

SSRs: 3%

a COMMON single base substitution between individuals (rare single base substitutions = mutations)

serve as biological markers

• in coding sequences of genes (alleles)

• in non-coding regions

• in the inter-genic regions between genes

• protect the ends of chromosomes from degradation

• with each round of replication, telomere lengths get shorter & shorter

1000-1700 copies [~10 kb] of the hexameric sequence TTAGGG

a reverse transcriptase (polymerase/enzyme - uses an RNA transcript) that makes telomeres

restores telomere length in the germ-line of each generation

tumor cells

enzyme rejuvenates the telomeres & circumvents the mitotic clock

the loss of 50-200 nucleotides from the ends of chromosomes every time a cell divides

when the telomeres are sufficiently shortened, the cell is no longer able to divide