Team:Tokyo Tech/Experiment/RNA thermometer assay

RNA thermometer assay

We have characterized previous parts.

　　

contents

1. Introduction

2. Summary of the Experiment

3. Results

4. Discussion

5. Materials and Methods

5.1. Construction

5.2. Assay Protocol

5.2.1. Protocol1

5.2.2. Protocol2

6. Reference


　　

1. Introduction

　　　　　　

ここにコピペ。

2. Summary of the Experiment

　　　　　　

ここにコピペ。

3. Results

　　　　　　

ここにコピペ。

4. Discussion

　　　　　　

ここにコピペ。

5. Materials and Methods

5.1. Construction

-Strain

　　　　　　

All the samples were DH5alpha strain.

-Plasmids

　　　　　　

Sample:J23119 promoter_RNA thermometer(FourU)_RFP(

Fig.3-8-何とか-1.


5.2. Assay Protocol

1. Prepare 2 over night cultures for each sample in 3 mL LB medium containing chloramphenicol (25 microg / mL) at 37 ºC for 12 h.
2. Dilute the overnight cultures to 1/100 in fresh LB medium (3 mL) containing chloramphenicol (25 microg / mL) (fresh culture).
3. Incubate the fresh cultures at 37℃ for 8 h.
4. Start preparing the flow cytometer 1 h before the end of incubation.
5. Measure the OD590 and adjust the volume of each sample to centrifuge so that the amount of pellet will be about the same for every sample.
6. Centrifuge the samples at 9000x g, 1 min , 4℃.
7. Remove the supernatants by using P100 pipette and suspend the samples with 1 mL of filtered PBS (phosphate-buffered saline).
8. Dispense all of each suspension into a disposable tube through a cell strainer.
9. Measure fluorescence intensity with flow cytometer.

6.. Reference

　　　　　　

ここにコピペ。