Lecture 2 Properties of DNA

Reference:Chapter 28: Biochemistry by Voet and Voet

C-value paradoxC-value paradox

Gene size, genome sizeGene size, genome size

Topology of DNA Topology of DNA

Denaturation and renaturation of DNADenaturation and renaturation of DNA

Lecture 2: Outline

gene sizes Largest known mammalian gene is

DMD gene 2.5 Mbp (0.1% of the genome)

Causes Duchenne’s muscular dystrophy (DMD)

characterized by rapid progression of muscle degeneration which occurs early in life.

‘scoliosis’

DMDMDDDMD gene maps to

chromosome Xp21DMD affects mainly malesMutations in DMD gene lead to non functional dystrophin protein

genome sizes

organism Number of base pairs (kb)

 

 

virusesLambda bacteriophage ( λ) 48.6

bacteriaEschericia coli 4,640

eukaryotesYeast 13,500Drosophila 165,000Human 3.3 x 106

Does size matter? C-value Does size matter? C-value paradoxparadox

mountain grasshopper Podisma pedestrisGenome size: 18 Gbp

protozoan Amoeba dubiaGenome size: 670Gbp

Boa constrictorGenome size: 2.1 Gbp

Homo sapiens sapiensGenome size: 3.2 Gbp

C value: DNA content of a haploid cellC value: DNA content of a haploid cell

CComparatiomparative genome ve genome sizessizes 

 

Why is there a discrepancy between genome size and genetic complexity?

C-value paradox

 

Due to the presence of repetitive (junk?) DNA

Repetitive DNA families constitute nearly one-half of genome (~45%)

Protein domains contribute to organism complexity

Topology of DNA

DNA supercoiling: coiling of a coil

Important feature in all chromosomes

Supercoiled DNA moves faster than relaxed DNA

Allows packing / unpacking of DNA

negatively supercoiled (right handed)•Results from under or unwinding•Important in DNA packing/unpacking e.g during replication/transcription

positively supercoiled (left handed)•Results from overwinding•Also packs DNA but difficult to unwind

Supercoiling topologySupercoiling topology

No supercoiling (left) to tightly supercoiled (right)

Analogy = phone cord

Supercoiling takes 2 forms Supercoiling takes 2 forms toroidal (DNA around histones) or toroidal (DNA around histones) or

interwound (bacterial chromosomes)interwound (bacterial chromosomes)

Supercoiled DNA needs to be Supercoiled DNA needs to be unwound /wound when necessary unwound /wound when necessary Unwound during transcription/replicationWound before cell division

Supercoils can be relaxed by Single strand nicks (topoisomerase I) Double strand breaks (toposiomerase II)

Topoisomerase I

Cause temporary single strand breaks in DNA Allows free rotation of helixActs by breaking phosphodiester linkage via tyrosine active site

Topoisomerase II (DNA gyrase)

ATP hydrolysis mediated double strand breakInhibitors are effective antibiotics and cancer chemotherapy agents

supercoiled

Relaxed circle

Full length linear

Why does a plasmid (circular DNA) that has never been cut give more than one band on a gel?

EBr

Denaturation and Denaturation and renaturation of DNArenaturation of DNA

Applications in polymerase chain reaction (PCR)

http://www.dnai.org/b/index.html

Denaturation of DNA

Also called melting

Occurs abruptly at certain temperatures

Tm – temp at which half the helical structure is lost

DNA melting curve

Tm varies according to the GC content

High GC content - high Tm

GC rich regions tend to be gene rich

Renaturation of DNA

Also called annealing

Occurs ~ 25oC below Tm

Property used in PCR and hybridisation techniques