导航：首页-->生物技术-->分子生物学技术指导（在线分子生物学技术方法）
- Genome-wide Gene
Expression Analysis (Richard Young Research
Group,Whitehead Institute for Biomedical Research)A genoe-wide gene
expression analysis using high-density oligonucleotide arrays
- Restriction Digests
of High Molecular Weight Yeast DNA (Donis Keller
Lab)
Restriction digests of YACs for mapping using rare cutting enzymes or
more conventional restriction endonuclease digestion.
- Preparation
of Yease Cell for ImmunoEletron Microscopy (Jon Mulholland, Botstein
Lab)
- Yeast Immunofluorescence
with Methanol/Acetone Dehydration (Botstein Lab)
- Yeast Immunofluorescence
(without methanol) (D. Amberg, Botstein Lab)
- Yeast
Rhodamine-Phalloidin/Calcofluor Staining (D. Amberg, Botstein
Lab)
- Yeast Cell
Cycle by Flow Cytometry (Forsburg Lab, The Salk
Institute for Biological Studies)
- Vectorette
PCR of Yeast DNA
(Botstein Lab)
- Yeast Actin Capture Assay
(D. Amberg)
- Yeast Preparation
for FACS Analysis (Gottschling
Lab)
- Replication Timing by Density Transfer (Fangman/Brewer lab)
- Replication Timing by Comparative Hybridization
(Fangman/Brewer lab)
- Replication Timing Using Transient
Hemimethylation (Fangman/Brewer
lab)
- ß-GAL Filter Assay (Breeden
Lab)
- Bgal
filter assay (Herskowitz Lab)
- Bgal plate overlay assay (Herskowitz Lab)
This assay is semiquantitative. Cells can even be
recovered from underneath the overlaid agar. The method could be used for any
enzyme with a colorimetric assay.
- Bgal liquid assay
(Herskowitz Lab)
- Pheromone Halo Assay (Dohlman Lab)
This is a bio-assay that measures the responsiveness of
cells to a factor pheromone. The assay is easy to conduct and the results are
usually unambiguous and highly reproducible.
- Liquid ß-gal Assays (Dohlman
Lab)
This protocol is used to quantitatively measure the pheromone response
over a range of pheromone concentrations.
- ß-gal Lift
Protocol (Dohlman Lab)
This method is used to screen for and examine
mutants with altered pheromone responses.
- Handling and using insertion
libraries (YGAC)
- Yeast Gene
Mutagenesis
- Protocols for shuttle mutagenesis/epitope-tagging of a yeast gene with
mTn-lacZ/LEU2 (Yale Yeast Analysis
Center)
mTn-lacZ/leu2 can easily be inserted at mutiple sites in a given
gene. The mutagenized DNA is then transformed into yeast, where it replaces the
chromosomal locus by homologous recombination. The transposon insertions create
a pool of insertion/disruption alleles. Insertions that generate in-frame fusion
of the coding region to lacZ can be used to monitor and quantify gene
expression, via assays for beta-gal activity. The fusion protein can also be
immunodetected using antibodies directed against beta-gal.
- Protocols for shuttle mutagenesis/epitope-tagging of a yeast gene with
mTn-3xHA/lacZ (Yale Yeast Analysis
Center)
Same protocol as above
- Mutagenizing a yeast gene with an mTn
(YGAC)
- UV Mutagenesis (Corbett
Lab)
- EMS
Mutagenesis (Herskowitz Lab)
- EMS
mutagenesis of yeast
(The Hahn Lab, The Fred Hutchinson Cancer Research Center and Howard Hughes
Medical Institute, Seattle)
- Generating a Temperature Sensitive Allele (Herskowitz Lab)
- Confirmation
PCR (Saccharomyces Genome Deletion Project)
To study the genetics
of yeast, yeast gene ORF is deleted by
PCR based strategy to generate mutated strain. Confirmation PCR is used to
verify if the yeast ORF
deletion is successful
- Introduction/General
strategy
Confirmation PCR primer design
Primer
organization in the 96-well plates
Oligonucleotide
concentrations
Quality
control procedures
Single
tube PCR protocol
96-well
format protocol
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