Oct. 03, 2008 six per page

Oct. 03, 2008 six per page - 1 Safety Common sense! Sign...

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Unformatted text preview: 1 Safety Common sense! Sign the safety sheet. Which and why? Which is on top? CAUTION: DO NOT HOLD THE FOCUS KNOBS ON BOTH SIDES SIMULTANEOUSLY. Ocular lenses Objective lenses Stage w/slide holder Focus Knobs Light Source & Field Diaphragm Aperture Diaphragm Hand Hold: Do NOT use! Poor design. 2 ! # $ % & ( ) * !+ ! !# !$ !% !! !( !& !, -./01 2/3456 (, 78014910 :3;861;<= !$, >?@05AB0 C.D083050 #, E;13;?F0 ?13<64C0DD 5.C41.F ), G.C:0CD01 603<64 !%, HI083050D $, JFF9=3C;4.1 *, 24;<0 !&, K3C.59F;1 60;: %, L30F: :3;861;<= !+, 2F3:0 6.F:01 !, L3C0 M.59D3C< NC.? &, G.C:0CD01 !!, G.;O3;F D4;<0 5.C41.FD !(, G.;1D0 M.59D3C< NC.? , G.C:0CD01 ;:@9D4=0C4 NC.? !#, >?@05AB0D CAUTION: Do not hold the focus knobs on both sides of the scope at the same time. CAUTION: Do not hold the focus knobs on both sides of the scope at the same time. CAUTION: Do not hold the focus knobs on both sides of the scope at the same time. Center field diaphragm CAUTION: Do not hold the focus knobs on both sides of the scope at the same time. Brightfield-unstained Brightfield-stained 50 m ? Fluorescence Fluorescence - confocal About 40 3 Which and why? How big? Which is on top? Gram + Gram - 1 m 2 m Astrographics.com E. Coli Yeast ? ? San Francisco Sourdough Starter 4 Eukaryotic cells 1 cm 0.1 mm 50 m 10 m Human cheek cells stained with methlylene blue. Tongue scrapping. Constructing recombinant DNA 1) Isolate DNA- source (foreign) DNA and vector DNA (source DNA has gene of interest: GOI) 3) Restriction digestions - generate smaller pieces 5) Ligation reaction - insert foreign DNA into vector DNA 7) Transformation - introduce recombinant DNA into host 5) Screen - look for transformed cells with gene of interest Screen: Amp R !-gal Objective: clone a Gene of Interest (G.O.I.) 5 Anneal & Ligate Isolate & Digest Transform & screen Weak Strong 6 Anneal & Ligate Isolate & Digest Transform & screen Restriction sequence for Hin d III 5 AAGCTT ---- AAGCTT ----- AAGCTT 3 3 TTCGAA ---- TTCGAA ----- TTCGAA 5 Piece 1 Piece 2 5 AAGCTT ---- AAGCTT ----- AAGCTT 3 3 TTCGAA ---- TTCGAA ----- TTCGAA 5 Piece 3 Piece 4 Sticky ends & annealing 5 A AGCT T ---- A AGCT T ----- A AGCT T 3 3 T TCGA A ---- T TCGA A ----- T TCGA A 5 Piece 1 Piece 2 Piece 3 Piece 4 Possible results? Ligation- sealing the ends! 5 A A-G-C-T- T----A 3 3 T- T-C-G-A A----T- T-C-G-A 5 Piece 1 Piece 2 5 A A-G-C-T- T----A 3 3 T- T-C-G-A A----T- T-C-G-A 5 5 A- A-G-C-T- T----A 3 3 T- T-C-G-A- A----T- T-C-G-A 5 Annealing- forming base pairs! Isolate & Digest Anneal & Ligate Anneal & Ligate Isolate & Digest Transform & screen 7 Purified " DNA A AGCTT---A AGCTT TTCGA A---TTCGA A Ligation reaction already performed for you R.E....
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Oct. 03, 2008 six per page - 1 Safety Common sense! Sign...

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