Sam:LabNotes/Microbione/2009-3-6: Difference between revisions
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>Sam Chiang No edit summary |
>Sam Chiang No edit summary |
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*Prepare four 1.5-mL tube, labeled and wrapped with foil. | *Prepare four 1.5-mL tube, labeled and wrapped with foil. | ||
*Set up as following exp design: | *Set up as following exp design: | ||
No-treat 100uL 200uL 400uL | |||
--------------------------- | |||
H2O | H2O 400 0 0 0 uL | ||
2X SYBR | 2X SYBR 0 100 200 400 uL | ||
E. | E. coli 1 1 1 1 uL | ||
--------------------------- | |||
*Mix by gently pipetting (p1000 pipettor). Incubate at dark place for 15 min. Agitate by vortexing for every 3~4 min. | *Mix by gently pipetting (p1000 pipettor). Incubate at dark place for 15 min. Agitate by vortexing for every 3~4 min. | ||
Line 32: | Line 32: | ||
*load 1 uL sample from each tube on the slide and place with cover slips. | *load 1 uL sample from each tube on the slide and place with cover slips. | ||
*Keep the slide in dark storage box for microscope visualizing. | *Keep the slide in dark storage box for microscope visualizing. | ||
==Results== |
Revision as of 23:51, 6 March 2009
=Bacteria labeling using SYBR gree
Objective
- Using SYBR to stand E. coli and confirm using inverted flourescent microscope
- Reference
Samples & Materials
- Bacteria: E.coli suspension in PBS (OD600=0.913 -> around 10^9 CFU/mL = 10^6 CFU/uL)
- SYBR 2X: Diluted from SYBR 50X stock solution
Procedures
- Dilute SYBR 50X to 2X with RNAse-free H2O: 40 uL of 50X SYBR + 960 uL H2O -> 2X SYBR, 1000 uL.
- Prepare four 1.5-mL tube, labeled and wrapped with foil.
- Set up as following exp design:
No-treat 100uL 200uL 400uL --------------------------- H2O 400 0 0 0 uL 2X SYBR 0 100 200 400 uL E. coli 1 1 1 1 uL ---------------------------
- Mix by gently pipetting (p1000 pipettor). Incubate at dark place for 15 min. Agitate by vortexing for every 3~4 min.
- After incubation, add H2O to 1000 mL for each tube, the new concentration for E. coli -> 1000 CFU/uL)
- Prepare two microscopy slide with labeling.
- load 1 uL sample from each tube on the slide and place with cover slips.
- Keep the slide in dark storage box for microscope visualizing.