Standard yeast media and protocols were used to manipulate yeast strains listed in Supplementary Table 1. The C-terminal tagging and deletion of yeast proteins was made by homologous recombination of respective genes with PCR cassettes previously described (Janke et al., 2004) or constructed in the lab. For microscopy, strains were grown to a log phase in a low fluorescence SD-Trp, -Ura medium (prepared from LoFlo YNB; Formedium) containing 1 M sorbitol where indicated. To compare behaviour of Sla2 force sensors, Sla2-mNeonGreen and Abp1-mScarlet-I at endocytic sites, 4 min movies were acquired with 1 s frame rate by filters set FF01-504/12 (excitation), FF520-Di02 (dicroic) and FF01-542/27 (emission) for mNeonGreen, and with 0.5 s frame rate by FF01-562/40 (excitation), FF605-Di02 (dichroic) and FF01-647/57 (emission; all Semrock) for mScarlet-I fluorescence.
Ff01 542 27
Manufactured by IDEX Corporation
The FF01-542/27 is a narrow bandpass optical filter manufactured by IDEX Corporation. It is designed to transmit a specific range of wavelengths while blocking all others. The filter's primary function is to selectively filter light, enabling precise control over the wavelengths used in various laboratory and research applications.
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Lab products found in correlation
Loflo ynb, by Formedium (1 mentions)
Ff01 562 40, by IDEX Corporation (1 mentions)
Ff605 di02, by IDEX Corporation (1 mentions)
Ff01 647 57, by IDEX Corporation (1 mentions)
Ff520 di02, by IDEX Corporation (1 mentions)
Ixon ultra 897, by Oxford Instruments (1 mentions)
Cfi plan apo vc 20, by Nikon (1 mentions)
Ff02 438 24, by IDEX Corporation (1 mentions)
2 protocols using ff01 542 27
1
Yeast Protein Tagging and Imaging
2
Monitoring ATP Dynamics in Arabidopsis
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Wild-type Arabidopsis and pgr5 mutants expressing recombinant Nano-lantern(ATP1) were grown in Murashige and Skoog medium, and the first or second leaves of 7-day-old plants were harvested. Detached leaves were injected with coelenterazine h as described above and then, to assess their luminescence, were immediately placed onto the stage of a microscope system (Ti-E (Nikon), which included a camera, iXon Ultra 897 (Andor Technology); objective lens, CFI Plan Apo VC 20× (NA = 0.75) (Nikon) and filters, FF02-438/24 (actinic light, Semrock), FF01-500/24 (mVenus excitation, Semrock) and FF01-542/27 (mVenus emission, Semrock).
The settings were as follows: exposure, 1 s; dead time, 300 ms; binning, 1; EMgain, Max. The luminescence in the leaves was measured in the presence and absence of actinic light ( max = 438 nm; 9.1 mW cm -2 ).
Rise and decay signals over a 7-s period were acquired immediately after dark-to-light or light-to-dark transitions and then were fit to single exponential curves. The luminescence rise and decay rates are reported herein as the half-life of each exponential curve.
We performed at least three replicates for each analysis.
The settings were as follows: exposure, 1 s; dead time, 300 ms; binning, 1; EMgain, Max. The luminescence in the leaves was measured in the presence and absence of actinic light ( max = 438 nm; 9.1 mW cm -2 ).
Rise and decay signals over a 7-s period were acquired immediately after dark-to-light or light-to-dark transitions and then were fit to single exponential curves. The luminescence rise and decay rates are reported herein as the half-life of each exponential curve.
We performed at least three replicates for each analysis.
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