实验动物技术

Preparation of the probe. Probes are prepared as Digoxigenin labelled RNA . The labelling mix as well as all antibodies are purchased from Boehringer All conditions and solutions should be totally RN ...

There are two kinds of contaminants on worm plates:1. Fungi: these contaminants can come from the plates or bacteria so it is best to leave plates out after seeding for a couple of days to make sure n ...

Stockkeeping1. MechanicsMost stocks can be successfully cultured by periodic mass transfer of adults to fresh food. Bottles or vials are tapped on the pounding pad to shake flies away from the plug th ...

Equipment flies frozen in 250 ml centrifuge bottlessieves: 106 355 600 and 850 mmpaintbrushcollection tubesfunneldry icejacket hat and gloves! Preparation 1. Freeze flies in 250 ml centrifuge bottle(s ...

Gloor et al. 1993 Genetics 135:81-95We have developed a simple method for the rapid and reproducible isolation of DNA from single flies for amplification by the polymerase chain reaction (PCR) (Saiki ...

.1 RNA Probe Preparation (see Note 1) 1. 1.5 mL microcentrifuge tubes or standard 96-well V-bottom microplates.2. RNAse free water.3. T7 T3 or SP6 RNA Polymerase (Fermentas Life Sciences Burling ...

ProcedureTransfer 1.5 ml of liquid culture of yeast grown for 20 - 24 h at 30°C in YPD (1% yeast extract 2% peptone 2% dextrose) into a microcentrifuge tube. Pellet cells by centrifugation at 20000 × ...

1.Inoculate a 5 ml culture with a single colony from a YAC-containing strain of yeast and grow until saturated. Determine the number of yeast cells per milliliter. The cell count should be roughly 1 x ...

Purpose: To perform restriction digests of YACs for mapping using rare cutting enzymes or more conventional restriction endonuclease digestion. Time required: 2-4 hours Special Reagents: dependent upo ...

Pick colonies into 0.5ml of SD-Leu (or other appropriate SD medium) Vortex for 1min Leave to grow O/N for 18-24h at 30°C 230-250rpm (best in 5ml bijou) Spin yeast culture at 13000rpm for 5 min (microf ...

Grow a 5 ml YPD O/N culture inoculated with a single yeast colony at 30 deg. Transfer culture to a small 13 x 100 glass tube. Spin down cells 2 min. in tabletop centrifuge. Pour off supernatant. Wash ...

Grow 100 ml yeast cells in desired media overnight to an A600 of ~1.0 (0.6 to 1.2 works well). For growth in minimal media 1 ml of a saturated overnight culture in minimal media (Synthetic dextrose (S ...

Yeast Media: Note: Synthetic complete medium can be prepared by adding media supplements (see below). Medium using 6.7 g yeast nitrogen base without amino acids (Difco # 0919-15) can also be prepared ...

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% 2-mercaptoethanol Incubate 10min on ice. Add 0.16 ...

-Use sterile technique and sterile solutions in steps 1 to 3.- 1. Using a saturated starter culture inoculate 25 to 30 ml of appropriate media in a 125 ml flask. ***Since it is often difficult to esti ...

Procedure pick one colony inoculate in 3 ml of the appropriate media grow at 30° overnight pellet the cells (5 min 5000g) wash 1X in sterile ddH2O re- suspend the pellet in 200 µl EthOH (optional: +2 ...

This is the preferred method for yeast RNA preparation use Gloves and RNAse free solutions throughout.1. Use a YPD overnight culture to innoculate fresh YPD media and grow cells at 30 degrees overnigh ...

Day 1Set up an overnight culture in 100 ml LMM broth or 100 ml terrific broth containing 100ul 100 mg/mlAmp Day 2Add 40-50 ml o/n culture to 1 lt terrific/K2K (see appendix) with 0.5 ml 100mg/ml Amp. ...

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 evening to get an of OD600 = 1 to 2 the next morning. ...

Transformation Protocol for Arabidopsis � Abbreviated Germinate seed in pots ↓ 4 weeks Streak bacteria onto YM/MinA ↓ 2-3 days 28°C Spray/dip bacterial suspension onto plants ↓ 1 day in box Let plants ...