Ciras 3 portable photosynthesis system

The CIRAS-3 portable photosynthesis system (PP SYSTEMS, MA, USA) was used to test the photosynthesis of each plant in the two treatments, including A (μmol CO₂ m^–2 s^–1), Ci (μmol CO₂ mol^–1), E (mmol H₂O m^–2 s^–1), and Gs (mmol H₂O m^–2 s^–1). Leaf photosynthesis was tested on the third youngest fully expanded leaf on each plant. All detection was conducted in accordance with the standard instruction of the meter. The detection location was in the controlled climate room, and the time was measured between 9:00 to 11:00 in the morning.

A CIRAS-3 portable photosynthesis system (PP Systems^®, Hitchin, UK), equipped with a PLC3 universal leaf cuvette, was used to determine photosynthetic capacity (A_max), transpiration (E), and water use efficiency (WUE) in response to increasing levels of photosynthetically active radiation (PAR), expressed as photosynthetic photon flux density (PPFD, 0–1600 µmol m⁻² s⁻¹). Measurements were conducted at 66 DODS and on three leaves per cultivar exposed to WW and DS treatments.

The photosynthetic capacity of the seedlings was determined every 5 days at 9.00 a.m. during the study period using a CIRAS-3 portable photosynthesis system (PP Systems, USA), according to Jia et al. [32 (link)]. Ten individually mature and fully expanded leaves were measured for each treatment. The chlorophyll fluorescence was detected before and after the treatments according to Jia et al. [31 (link)], using an Open FluorCam FC 800-O imaging fluorometer (Photon Systems Instruments), and analyzed with Fluorcam7 software (PSI, Brno, Czech Republic).

Gas exchange assessments consisted of non-destructive analyses of fifteen diagnostic maize leaves using a using a Portable Infrared Gas Analyzer CIRAS-3 Portable Photosynthesis System (PP Systems Inc., Amesbury, MA, United States). The following parameters were determined on the diagnostic leaves of maize: net photosynthetic rate expressed as area (A; μmol CO₂ m^–2 s^–1); stomatal conductance (gs; mol H₂O m^–2 s^–1); internal CO₂ concentration in the substomatal chamber (ic; mmol CO₂ mol^–1 air); leaf transpiration (E; mmol H₂O m^–2 s^–1); and water use efficiency [WUE; μmol CO₂ (mmol H₂O)^–1], calculated by the A/E ratio. Readings began after the air temperature in the chamber was adjusted to 28°C, with 380 ppm CO₂ and 1,000 μmol m^–2 s^–1 of photosynthetically active radiation (PAR) supplied by LED lamps. The measurements were performed between 8:00 and 10:00 a.m. The minimum equilibration time before performing the reading was 3 min.

Thirty mutant and WT plants were grown in the greenhouse. At the fruit setting stage (seventy-day-old), the vine length, width of the stem, number of nodes and internode length, and leaf area (the fourth functional leaf from the top) were measured. Each parameter was determined from 15 biological repeats. Phenotypes of the cucumber plants were recorded using an optical camera (D7100, Nikon, Japan).
We measured chlorophyll content and several photosynthetic parameters in mutant and WT plants. Leaf gas exchange was measured with a Ciras-3 Portable Photosynthesis System (PP-Systems company, Amesbury, MA, USA) on the fourth functional leaves from the top at the fruit setting stage under 1000 μmol·m⁻²·s⁻¹ PPFD at a controlled CO₂ supply (400 mmol CO₂ mol⁻¹ air). Parameters measured included: net photosynthetic rate (Pn), stomatal conductance (gs), intercellular CO₂ concentration (Ci), and transpiration rate (Tr). These leaf samples were also used for chlorophyll measurement using a mixed extracting solution (acetone:alcohol:distilled water = 4.5:4.5:1) at room temperature for 24 h and then measured with a Bio-Rad SmartSpec Plus spectrophotometer at 663 nm, 645 nm, and 470 nm, respectively, following Tang et al. [88 (link)]. Each parameter was determined from 5 biological repeats.

The water potential (Ψ_w, MPa) was measured with a Schölander pressure pump (model PMS-1000, PMS Instruments, Corvallis, OR, United States). Stomatal conductance (g_s, mmol H₂O m⁻²s⁻¹), the sub-stomatal concentration of CO₂ (C_i), photosynthetic rate (A, μmol CO₂ m⁻²s⁻¹), transpiration (E, mmol H₂O m⁻²s⁻¹), water use efficiency (WUE, μmol CO₂ mmol⁻¹H₂O) and leaf temperature through infrared Thermometry (Tleaf, °C), were determined with a CIRAS-3 portable photosynthesis system (PP Systems, Amesbury MA). The measurements were recorded under saturating light conditions (1,500 μmol quanta m⁻² s⁻¹), with a temperature of 25°C, and ambient CO₂ concentration of 400 mol⁻¹ CO₂ and a relative humidity of ~55%. Chlorophyll fluorescence indices (i.e., Fv/Fm and Quantum yield) were measured with a portable pulse-amplitude modulated chlorophyll fluorometer (PAM-2100, Heinz Walz, Effeltrich, Germany). These measurements of the photosystem II efficiency were performed once the plants were adapted to darkness for 30 min, on the same leaves where stomatal conductance and photosynthesis were determined. All measures were performed on the third youngest full-developed leaf of each plant, analyzing a total of five plants per variety.

These were measured at 20 days after salt treatment and selected the seventh of the full-expansion function leaves from bottom to top. Photosynthetic parameters in fully expanded leaves, including Pn, WUE, GS, and Tr were determined within the time period of 8:30 am to 10:30 am using a CIRAS-3 Portable Photosynthesis System (CIRAS-3; PP Systems, United States).