其他模型

I. SUMMARY The yeast two-hybrid system provides a relatively straight forward approach to understanding protein function. Section II outlines the basic components of ...

This Protocol allows You to prepare Frozen Yeast cells that are competent for transformation after thawingAfter Dohmen et al. (1991) Yeast 7: 691-692. See Schiestl et al (1993) The ability to make yea ...

Large-scale yeast transformationThe following protocols were developed by Petra Ross-Macdonald for Mike Snyder's LacZ fusion project at Yale University. Yeast Cell PreparationThis protocol merges a ty ...

Multiwell Transformation ProtocolLabor-saving implementsMultichannel pipettors--1 to 25 µl and 20 to 200 µl volume (Brinkmann 12-channel). Sterile 96-well assay or tissue culture plate with lid (such ...

Typical transformation resultsThe following transformations were performed at the same time using the multiwell transformation protocol. Similar amounts of PCR product were used for each. The pictures ...

Efficient Transformation of Yeast - LiAc MethodDifferent strains transform at different rates with different methods. Some strains transform much better using the electroporation method. Other ones ar ...

Note: Please citeAgatep R. R.D. Kirkpatrick D.L. Parchaliuk R.A. Woods and R.D. Gietz (1998) Transformation of Saccharomyces cerevisiae by the lithium acetate/single-stranded carrier DNA/polyethylene ...

Yeast Transformationinoculate 5-10 ml culture with single colony and grow ON measure OD600nm and dilute to OD600nm = 0.3 in 50 ml grow for 2-4 hrs spin wash cells with 25 ml H2O spin resuspend in 2 ml ...

Electro-Transformation of YeastDavid AmbergGrow cells to 1X10E8 or OD600 of 1.2-1.3. Spin cells at 5000rpm for 5 min and wash pellets in an equal volume of ice cold water. Wash in 1/2 volume cold wate ...

Yeast Transformation Using Frozen Competent Cells and Single-stranded DNA as a CarrierKatherine KolorReferences: Schiestl and Gietz (1989) Curr. Genetics 16:339-346 Gietz et al (1995) Yeast 11:355-360 ...

Yeast Genomic DNA IsolationDavid AmbergGrow 100 ml culture to about 1x10 to the 8th Spin 4 x 15 mls of culture down about 3k x 3 min. Resuspend entire pool in about 6 mls ddH2O aliqoute into 1.5ml mic ...

Yeast transformation using lithium acetate (rapid method)Steve HahnLast Modified December 1997This method works well when transforming with a plasmid and is rapid. (for highest efficiency transformati ...

Rhodamine-Phalloidin/Calcofluor StainingDavid AmbergGrow 50 mls of yeast to 5x10E6. Add formadehyde to the media to 4% (33 mls. of 10%). Fix in media at temperature for 10 min. Spin down cells 2-3 K f ...

Yeast Colony PCRAkada et al. Biotechniques 28:668-674 (April 2000)MATERIALS0.25% SDS10X Colony PCR Buffer: 0.125 M Tris-HCl pH 8.50.5625 M KCl25 mM MgCl210 mM dNTP's20% Triton X-100Taq polymeraseTwo G ...

Gene Disruption via PCR 14:115-132 (1998).Reaction mix:5 µl 10X Taq Buffer5 µl 25 mM MgCl22 µl 10 mM dNTP's10-100 ng template DNA25 pmols of each primer0.5 µl Taq polymerase (2 Units)_________________ ...

Quick Yeast DNA Prep: Isolation of Total DNA (genomic and plasmid) Linda Hoskins Hahn Lab 10/16/98Grow a 5 ml YPD O/N culture inoculated with a single yeast colony at 30 deg. Transfer culture to a sma ...

Genomic DNA Miniprepgrow 5 ml cells over night at 30 °C spin wash once with 1 ml H2O resuspend in 500 µl lysis buffer add acid washed glass beads to 1.25 ml vortex 2 min recover liquid phase with blue ...

Yeast Genomic DNA Prep Linda Hoskins/Hahn lab Aug 18 1997(modified from Philippsen 1991)Grow 10ml YPD cultures o/n. Figure out cell density; inoculate 30 ml YPD and grow o/n so that cell density is ap ...

Pick a medium size yeast colony (~3 mm dia) and transfer to 200 ul of lysis buffer. Add an equal volume of glass beads (0.45 mm dia). Mix on vortex at top speed for 1 min. Add 200 ul of phenol/CHCl3 ...

Yeast Miniprepfrom Robzyk and Kassir Nucleic Acid Res. (1992) 20:3790Materials:• 10 ml overnight culture yeast in drop out media (must be very dense)• STET (8% sucrose 50 mM Tris pH 8 50 mM EDTA 5% Tr ...