genetic engineering (4)

DNA replication
semiconservative
1.helix-destabilizing protein-helicase
2.single strand DNA binding protein (SBP, DBP)
3. DNA gyrase or topoisomerase-replication fork
4.primase-synthesize small RNA primer
5.DNA polymerase
E coli- pol3 leading strand 5'-->3', lagging strand small fragment (Ogazaki fragment), pol I 5'->3' exonuclease)
polI (C-terminal -polymerase, 3'-5' exonuclease -Klenov fragment, short N-terminal, 5'-3' exonuclease) -proofreading
Mammals-alpha-nuclear-polymerase, beta-nuclear-repair, delta -nuclear similar alpha, gamma-mitochondria-mitochondrial DNAreplication
6. DNA ligase
bidirectional
E coli- 1 replicon
Yeast-500 replicon
Drosophila-3500 replicon
Toad-15000 replicon
Mice 25000 replicon
plant -35000 replicon

Model
1. Theta model-circular DNA-bacteria, mitochondria
2. Sigma model-rolling circle model: phage lambda circular dsDNA-nick, unwheel, produce linear DNA-concatamer, catenated DNA, phage M13 circular ssDNA --> circular dsDNA--> linear ssDNA-->circular ssDNA
3. Y-shaped model-phage T7 linear dsDNA, higher organisms- chromosome

telomerase - terminal deoxynucleotidyltransferase - repeat CCCTAA---> inverted to complementary
found in egg and sperm only
so shorten of chromosome each replication 50-60 times --.senescence

Genetic engineering (3)

Structure and type of RNA
RNA - 5X DNA

transfer RNA (tRNA)80-100 type
5S ribosomal RNA 1-2 type
5.8 Sribosomal RNA 1 type
16 S ribosomal RNA 1 type
23 S ribosomal RNA 1 type
18 S ribosomal RNA 1 type
28 S ribosomal RNA 1 type
messenger RNA (mRNA) many thousands type
heterogeneous nuclear RNA (hnRNA) many thousands type
small cytoplasmic RNA (scRNA)
small nuclear RNA (snRNA)

Size, organization, complexity of Genomes
Size and organization of genomes
1000 bp to 10000 mbp
dsDNA except virus: ssDNA, dsDNA, ssRNA, dsRNA, linear or circular, single or multiple
bacteria-circular dsDNA+protein
eukaryote-linear dsDNA (multile)+histone, non histone protein

left hand super helix 1.8 rounds, 145 bpDNA(2nm)+2H2A, 2H2B, 2H3, 2H4- nucleosome core particle(10nm)--> solenoid(30nm)-->filament(300nm)-->supercoiled(700nm)-->chromosome (metaphase 1400nm)
60 bp linker DNA+H1

heterochromatin-inactive-centromere, telomere
active chromatin-10%

packing ratio=DNA length/Final length average 1000-2000

Complexity of genome
unique sequence, repetitive sequence
slow component 1 copies-45% of genome
intermediate component 350 copies-30% of genome
fast component 500000 copies-25% of genome

satellite, minisatellite, microsatellite

Important of DNA
DNA replication
DNA transcription
Complexity of genome