chapter 10. geneticswebbuild.knu.ac.kr/~app-mic/resources/lecture... · 2017-11-16 ·...

1. Bacterial DNA1) Bacterial chromosome

- most of genetic information is located within a single chromosome

- double helix in closed loop

- exists feely in cytoplasm without any protein support

- 1.5 mm long, 1500 times the length of the bacterium

- a number of small loops attached at anchorage point to be packed into small cell

- 4,000 genes for E. coli, 7 gene for virus, 50,000 gene for human 22 pair

Chapter 10. Genetics

☞ Nucleic acid (DNA structure)

- nucleotide의 중합에 의한 polynucleotide 구조

- nucleotide : pentose + base + phosphate group

- DNA, RNA


☞ Chromosome replication (복제)

- first DNA anchors to a point on cell membrane -> double helix unwind ->

DNA polymerase synthesis new strand (5’->3’)- closed loop is unwound by enzyme at the origin of replication

- semi-conservative(반보존적) method ->binary fission


☞ Replication process and Enzymes

- primase(primer 합성), helicase, DNA polymerase, ligase


2) Plasmid

- a number of closed loops of DNA

- contain 2% of genetic information in bacterium

- multiply independently of the chromosome

- shown mostly in Gram- bacterium

- not essential for bacterial life but confer selective advantage

-> resistance to antibiotics, produce toxin (bacteriocin) to struggle for life


2. Gene mutation (유전자 돌연변이)- change of genetic information by mutation and recombination(조합)- mutation : permanent change in an organism’s DNA

-> disruption of nucleotide sequence in a gene

-> loss of significant part of the gene

-> cause of change in mRNA sequence -> change in a.a sequence


1) Cause of mutation

- regularly spontaneous change -> once per 109 replication

-> appearance of antibiotic-resistant bacteria

- mutagens (돌연변이원) :

-> UV : produce thymine dimer

-> chemicals : urea, benzopyrene(매연) -> deletion or insertion of nucleotide

-> nitric acid

- transposon :

-> small DNA segment moving from one position to another in chromosome


Thymine dimerThymine dimer

☞ mutation by nitric acid☞ mutation by nitric acid


3. Gene recombination (유전자 재조합)- gene transfer (gene recombination) can occur between bacteria by three methods

-> conjugation, transduction, transformation

- gene recombination : genetic alterations by acquisition of DNA from others

1) conjugation (접합)- two live cells come together

- donor cell transfers some of its genetic material to a recipient cell

- F+ cell : donor cell vs F- cell : recipient cell F : fertility

- F factor : plasmid of the donor cell

-> contains 20 genes encoding enzymes and protein for conjugation

- F pili (sex pili) : a channel for gene transfer -> conjugation bridge (접합다리)


- Shigella, Escherichia, Salmonella …- gene of bacterial chromosome can be

transferred by transposon

-> transposon -> F factor -> conjugation

☞ Hfr (high frequency of recombination)- F factor attached to the chromosome

- Hfr donor cell has sex pili

- part of DNA enter the recipient cell

세균접합방법의 비교세균접합방법의 비교

2) transduction (형질도입)- gene transfer occurs with the assistance of bacterial virus (bacteriophage)

- happens very rarely in nature


☞ Generalized transduction

- during the lytic cycle, newly forming phage pick up fragment of bacterial DNA

- transfer gene during second lysogeny

☞ specialized transduction

- during lysogeny -> lytic process,

bacterial gene can be transferred to second cell

3) transformation (형질전환)- acquire genes from surrounding

environment

- direct uptake of DNA fragments

by a recipient cell

- competence factor : protein

helping gene uptake

What is happening because of microbial genetic recombination?


- antibiotic resistance of pathogenic microbes

- Staphylococcus aureus -> initiate disease when penetrated the skin

-> 패혈증, 간염, 뇌막염 …-> MRSA (multidrug-resistant Staphylococcus aureus) 출현

Microbial genetic recombination is devilish monster?

4. Genetic Engineering (유전공학)

- how to control those microbial genetic recombination ?

- during 1970s, alteration of bacterial DNA became possible

-> cutting, splicing of DNA, -> removing, inserting of genes

- 1960s, endonuclease was isolated -> react with nucleic acids within the cell

-> called restriction enzyme : act at restricted locations in nucleic acid

-> palindrome structure

-> biochemical scissor


- 1971, Daniel Nathans split DNA of SV40(simian virus) using bacterial enzyme

- restriction enzyme works on the same restriction site regardless of sources of DNA

-> plant, animal, bacterium, virus

- use restriction enzyme to alter DNA in a test tube ?

- 1971, Paul Berg -> constructed the first recombinant DNA

-> sliced viral DNA into E. coli chromosome

-> used blunt end

- 1971, Herbert Boyer isolated EcoRI nicking staggered end on the chromosome

-> stronger complementary pairing than blunt end

-> still not strong enough to hold the ends together forever

- DNA ligase is necessary to join the backbones of DNA strand

- 1972, Stanley Cohen used bacterial plasmid resulting in successful transformation

-> calcium shock (suspension in cold CaCl2 -> rapid heating to 42℃)

- first recombinant protein of toad cell protein in E. coli

-> beginning of the era of genetic engineering


Construction of a recombinant DNA molecule

chimera

BiotechnologyBiotechnology

Genetic engineeringGenetic engineering

chapter 10. geneticswebbuild.knu.ac.kr/~app-mic/resources/lecture... · 2017-11-16 ·...

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