By Canadian Mathematical Society
Read Online or Download Crux Mathematicorum with Mathematical Mayhem - Volume 34 issue 1-8, 2008 PDF
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Extra info for Crux Mathematicorum with Mathematical Mayhem - Volume 34 issue 1-8, 2008
Sample text
Check the primers for complementarity. 6. 95 μl H2 O. 7. Undertake PCR at 94◦ C 2 min, then 30 cycles of 94◦ C for 1 min, 60◦ C for 2 min, 72◦ C for 7 min and 72◦ C 10 min. See Note 2. 8. Purify the PCR products using Qiaquick PCR purification kit. 9. 2. 10. Transfer PCR products into pENTR/D-TOPO vectors as outlined in the kit. 11. 5-ml tube and store at –70◦ C. 12. PCR or sequence to confirm entry of PCR products into pENTR/D-TOPO using the M13 forward primer and the reverse ORF-specific primer.
Cell addition: 5. Count Confluent HEK293T cells with a haemocytometer, add 1×107 HEK293T cells to a T75 flask and make up to 15 ml with culture medium. Incubate at 37◦ C, 5% CO2 for 24 h. 6. Before cell addition, vortex and incubate 16 μl enhancer and 150 μl EC buffer per slide at RT for 5 min. 7. Add 25 μl effectene and vortex. Pipette the transfection reagent solution onto one end of the slide. Cut a plastic piece of film; cut to the size of the slide and carefully place onto the slide. Incubate at RT for 20 min.
Novel Fluorescent Transcriptional Reporter for Cell-Based Microarray Assays 49 8. Slides were rinsed in dH2 O and stored dry at room temperature in the dark. 6. Microscopic Imaging and Quantitation 1. Imaging of the ELF-97-stained cells was completed immediately following the staining procedure. 2. Cells were viewed using an inverted fluorescent microscope using a cube specific for the 365/530 nm wavelength of ELF-97. 3. A Cool Snap HQ camera was used to capture the images. Images taken for EGFP are 10-fold higher in length of time than those of the ELF-97.