7.8.Exp.methods.v4 - DNA methods summary 1 Rest r i ct i on enzym cut at speci f i c DNA si t es N es 2 Vect or s al l ow genes t o be cl oned and

Info iconThis preview shows pages 1–11. Sign up to view the full content.

View Full Document Right Arrow Icon
Background image of page 1

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
DNA methods summary 1. Rest r i ct i on enzymes cut at speci f i c DNA si t es. ( N) 2. Vect or s al l ow genes t o be “cl oned” and pr ot ei ns “expr essed”. ( N) 3. Gel el ect r ophor esi s separ at es DNA on t he basi s of si ze. 1. DNAs can be synt hesi zed ( up t o ~ 100 bases commer ci al l y) . ( N) 1. PCR ampl i f i es any t ar get DNA sequence. ( N) 1. Genes and genomes can be sequenced by chai n t er m i nat i on. ( N) 2. Ol i gonucl eot i des can be used t o change bases by si t e- di r ect ed mut agenesi s ”. ( N) 1. “Sout her n” bl ot t i ng det ect s sequences by hybr i di zat i on. 1. Genes can be knocked out ( del et ed) or r epl aced i n pr okar yot es and eukar yot es. ( N) 1. Mi cr oar r ays det ect gene expr essi on pat t er ns over t he genome.
Background image of page 2
Restriction enzymes cut DNA at specific sites
Background image of page 3

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Restriction enzymes cut DNA at specific sites Pal i ndr ome
Background image of page 4
Restriction enzymes cut DNA at specific sites Pal i ndr ome not e, I di ssent . A f ast never pr event s a f at ness. I di et on
Background image of page 5

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Restriction enzymes cut DNA at specific sites t ypes of ends: 5’ over hang, bl unt and 3’ over hang ognat e met hyl t r ansf er ases pr ot ect host genome f r om di gest i Rest r i ct i on- modi f i cat i on syst ems degr ade “f or ei gn” DNA.
Background image of page 6
Average frequency of restriction sites in “random” DNA sequences e aver age occur r ence of each sequence = 1/ 4 n , er e n = t he si t e l engt h and al l bases ar e equal l y r epr esent Si t e si ze 4 6 8 Aver age f r equency 1/ 256 1/ 4, 096 1/ 65, 536 ( 1/ 4 x 1/ 4 x 1/ 4 x 1/ 4)
Background image of page 7

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Lots of different recognition sites known Cor e f our bases Fl anki ng bases None A- - - - T C- - - - G G- - - - C T- - - - A
Background image of page 8
A simple cloning procedure 1. Cut “i nser t ” and “vect or ” DNA wi t h a r est r i ct i on enzyme 1. Mi x and j oi n ends wi t h DNA l i gase. The ends shoul d mat ch f or ef f i ci ent l i gat i on.
Background image of page 9

Info iconThis preview has intentionally blurred sections. Sign up to view the full version.

View Full DocumentRight Arrow Icon
Cloning without DNA ligase 1. Pr epar e open vect or and i nser t wi t h t he same l ong “st i cky” ends + Pol I Kl enow f r agment + dATP 1. Mi x and l et t he ends anneal . 1.
Background image of page 10
Image of page 11
This is the end of the preview. Sign up to access the rest of the document.

This note was uploaded on 03/08/2010 for the course MCB 110 taught by Professor Alber,zhou,nogales during the Fall '07 term at University of California, Berkeley.

Page1 / 31

7.8.Exp.methods.v4 - DNA methods summary 1 Rest r i ct i on enzym cut at speci f i c DNA si t es N es 2 Vect or s al l ow genes t o be cl oned and

This preview shows document pages 1 - 11. Sign up to view the full document.

View Full Document Right Arrow Icon
Ask a homework question - tutors are online