YeastGeneknockoutusingOligo／PCR
Universal primers for gene knock-out using dominant drug markers: Kan, Clonat, and Hygromisin-B. Forward primer: 5’ TCAGGGGCATGATGTGACT 3’Reverse primer: 5’ AGCTCGTTTTCGACACTGGAT 3’Plasmids for Selectable MarkersPlasmid for amplifying Kan:pKan-GenMX4.seq (GCK), product size: ~1.4kb.Plasmid for amplifying Clonat:pAG25-ClonatMX4.seq (GCK), product size: ~1.2kb.Plasmid for amplifying HB:pAG32-hphMX4.seq (GCK), prod......閱讀全文
Yeast－Gene－knockout－using－Oligo/PCR
Universal primers for gene knock-out using dominant drug markers: Kan, Clonat, and Hygromisin-B.?Forward primer: 5’ TCAGGGGCATGATGTGACT 3’Reverse prim
Targeted－Gene－Replacement－in－Fungi－Using－a－SplitMarker－Approach
Targeted gene replacement is one of the primary strategies for functional characterization of fungal genes and several methods have been developed
酵母遺傳學技術
Genome-wide Gene Expression Analysis?(Richard Young Research Group,Whitehead Institute for Biomedical Research)A genoe-wide gene expression analysis u
轉基因——基因標靶
Gene Targeting Outline?(University of Michigan Transgenic Animal Model Core)This is a brief outline of the steps necessary to produce mice with a muta
Yeast－Media
YEPD (non-selection)-1% yeast extract-2% peptone-1.5% agar (if needed for plates)After autoclaving add glucose to 2% by adding 100 ml of 20% solution
其它PCR方法
·?????????Standard PCR Protocol?(Molecular Biology Techniques Manual)The followings are described in detailRecommended Reagent ConcentrationsRecommend
Using－GenBank
GenBank(R) is a comprehensive database of publicly available DNA sequences for more than 205,000 named organisms and for more than 60,000 within t
Dropout－plates－for－yeast
Materials(Solutions are all available from the media room)200ml bottle of 2x SD200ml bottle of 4% agar -- make sure to sign it out40% glucoseCSM minus
Yeast－DNA－Prep
Protocolgrow up yeast culture to appropriate density (near saturation)spin 1.5 mls of culture for 1 min in microfuge and aspirate off supernatantresus
yeast:Assaying－mating
SetupYou have yeast strains that are deficient in mating (eg Ste12 knockouts) and would like to test whether transforming them with a plasmid that con
Yeast－Lysates－for－Westerns
Cells are grown for 2-3 days as 1.5ml prep. under selection for the plasmid of interest. Spin cells down 2.6K for 5min.Resuspend in 1ml 0.25m NaOH/1%
Preserving－yeast－cultures
Short term storageYeast cultures are stable for 1-2 weeks when refrigerated. Petri dishes should be sealed or in plastic bags.Medium term storageYeast
Yeast－Nuclei－Isolation
This method gives yeast nuclei which look nearly purified microscopically. Nuclei isolated in this way do not give active transcription extracts when
Modified－Yeast－Transformation
Inoculate cells from an overnight culture into 50 ml YEPD and incubate at 30°C with shaking. Typically, add 0.1 to 0.2 ml saturated culture in the eve
Fast－Yeast－Transformation
Protocol: Fast yeast transformationAdd 50 μl carrier DNA to a 1.5 ml tube.scrap cells from plate and add to the carrier DNA.Add in the following order
Using－a－Counting－Chamber
Using a Counting ChamberFor microbiology, cell culture, and many applications that require use of suspensions of cells it is necessary to determine ce
無創血壓計應用論文：動物用血壓計（一）
Androgen Receptor Gene Knockout Male Mice Exhibit Impaired Cardiac Growth and Exacerbation of Angiotensin II-induced Cardiac Fibrosis?【摘要】? Androg
Yeast－Genomic－DNA－Prep
Grow 10ml YPD cultures o/n. Figure out cell density; inoculate 30 ml YPD and grow o/n so that cell density is approximately 2 x 108cells/ml the next m
Endy:Yeast－Colony－PCR
MethodUsing sterile pipette tips, transfer a 1 mm colony into 50 uL of 60 U/ml Zymolyase3 uL of 1 U/mL Zymolyase stock solution47 uL of waterIncubate
Decontamination－of－cells－from－the－yeast
I???? Destroy yeast1.???? Aspirate medium and wash cell in PBS.2.???? Incubate cells at 37oC for 5 min in non-diluted antibiotic-antimycotic.3.???? In
Live－Cell－Imaging－of－Yeast
Live Cell Imaging of YeastDaniel R. Rines, Dominik Thomann, Jonas F. Dorn, Paul Goodwin and Peter K. SorgerINTRODUCTIONThe development of cloning vect
Blackburn:Yeast－Colony－PCR
OverviewThis is a quick and easy yeast colony PCR protocol that does not require zymolyase step.Updated Protocol:?Blackburn Lab: Quick and Easy Yeast
Plasmid－isolation－from－yeast
Pick colonies into 0.5ml of SD-Leu (or other appropriate SD medium)Vortex for 1minLeave to grow O/N for 18-24h at 30°C, 230-250rpm (best in 5ml bijou)
Yeast－Media,－Solutions－and－Stocks
Yeast Media:Note: Synthetic complete medium can be prepared by adding media supplements (see below).Medium using 6.7 g yeast nitrogen base without ami
人工轉錄因子的部件——人類鋅指結構1
Human zinc fingers as building blocks in the construction of artificial transcription factorsKwang-Hee Bae1, 4, Young Do Kwon1, 2, 4, Hyun-Chul Shin1,
Yeast基因文庫的分類和選擇
文庫種類Dharmacon酵母資源包括多個酵母基因組文庫，包括ORF文庫、基因敲除（Knock Out）菌株、蛋白質相互作用文庫、突變菌株和各種篩選文庫等。除此之外，Dharmacon 針對Saccharomyces cerevisiae 研究領域提供了Zoonome siRNA 文庫。酵母
Deglycosylation－of－Glycoproteins－Using－Endoglycosidases
T.H. Plummer, Jr. and A.L. Tarentino, Department of Biochemistry, New York State Department of Health, Wadsworth Center for Laboratories and Research,
ChIP－using－plant－samples
實驗概要?The ?immunoprecipitation (IP) of cross-linked chromatin with antibodies ?specific for certain histone modifications (chromatin ?immunoprecipitati
A－Method－for－Assaying－Deubiquitinating－Enzymes1
AbstractA general method for the assay of deubiquitinating enzymes was described in detail using?125I-labeled ubiquitin-fused αNH-MHISPPEPESEEEEEHYC (
Adiponectin－Replenishment－Ameliorates－ObesityRelated－Hypertension（二）
Methods??? Animal and Animal Treatment??? KKAy male mice were purchased from Japan CLEA (Tokyo, Japan). This strain is a cross between black KK fema