Intensilight c hgfi 130w lamp

Neurospora crassa strains of the same mating type, which individually contain GFP and RFP, were cultivated in VMM + S slants at 25°C for 7 days to get enough macroconidia. The spores of each strain were harvested and washed with 1 M sorbitol. Then they were counted under a microscope, and their final concentrations were made to 6.5 × 10⁷ spores/ml. One hundred μl of each conidial sample from the GFP strains were individually mixed with the same amount of conidia from the LSP-1:RFP strain. They were incubated at 25°C for 30 min and then were pipetted into VMM + S slants. The slants were incubated at 25°C in light for 5 to 6 days. During the time the spores of both fluorescence were germinating, fusing, and developing into strains with dual fluorescence proteins. The conidial spores from each dual fluorescence strain were inoculated onto VMM + S thin agar plates and were cultivated briefly or were directly checked for fluorescence under the fluorescence microscope (ECLIPSE Ci system plus INTENSILIGHT C-HGFI 130w lamp, Nikon).

Fungi of different genotypes were incubated on thin agar plates before microscopy.
Microscope analysis was performed with a fluorescence microscope (ECLIPSE Ci system plus INTENSILIGHT C-HGFI 130 w lamp, Nikon). 40 × /NA0.75 (plan fluor), 100 × /NA1.30 oil (plan fluor) objective lenses were used to acquire images. Immersion oil (Type N, Nikon) was used with the oil objective lens for observation and image acquisition. Epi-Fluorescence filter G-2E/C (TRITC), EX 540/25, DM 565, BA 605/55 was used for red fluorescence. A GigE camera (DFK 23U274, Imaging Source) was used to capture photos. The acquisition software was NIS elements D basic (Nikon). Images were captured at room temperature (22–25°C).
Confocal fluorescence analysis was performed using a confocal laser scanning microscope (Leica, TCS SP5) and images were captured with the Leica LAS AF Lite Software. GFP was excited with 488 nm light and emission was detected at 500–550 nm, while RFP was excited with 543 nm light and emission was detected at 590–610 nm.
During each microscope experiment, including CLSM, a blank control of WT was set to make sure there was no auto fluorescence at the experimental settings.