Philodendron plant tissue culture
Philodendron is suitable for use as table ornamental plants, hanging baskets, totems or floor plants. Laboratory culture is a method of asexual reproduction under aseptic conditions that uses the ability of each plant cell to produce the same genetic material. Production of ornamental plants has used this technique to propagate plants with a high degree of uniformity and health. Most commercial plants of the genus Philodendron are propagated by in vitro tissue culture.
Philodendron is the second largest genus of the family Araceae , with 480 exclusively Neotropical species. The morphological variability that exists among the different species, the recently developed cultivars, and their growth habit makes philodendron cultivars suitable for use as ornamental desk plants, hanging baskets, totems, or floor plants.
Philodendron plant tissue culture
In vitro culture is an asexual reproduction technique under aseptic conditions, which exploits the ability of each plant cell to generate genetically identical individuals. Ornamental plant production has used this technique to multiply plants with a high degree of uniformity and health. Most commercialized plants of the genus Philodendron are propagated by in vitro tissue culture.
To initiate in vitro cultures, there is a serious problem with contaminants, as they compete advantageously with plants for the nutrients in the culture medium and end up killing the tissues . Therefore, it is necessary to develop efficient explant disinfection protocols that eliminate exogenous microorganisms
In this sense, the treatment of mother plants with fungicides and bactericides before in vitro establishment and during the disinfection process of philodendron explants is effective in achieving asepsis, since pathogens hosted in the plant organs are prevented and eliminated.
established the aseptic culture of three Philodendron cultivars by spraying the mother plants with Ridomil MZ® and Mancozeb® one week prior to explant excision and disinfested them with 70% ethanol for one min followed by a 1% (v/v) sodium hypochlorite solution for 20 min . achieved the aseptic establishment of Aglaonema sp. by soaking the explants in Antracol® for 30 min, then in 70% ethanol for two min and finally in 20 and 50% Clorox® for ten min.
Recently, the application of silver nanoparticles (AgNPs) in plant tissue culture has increased due to their ability to eliminate microorganisms that affect in vitro cultures . Explant disinfection protocols during in vitro establishment with AgNPs produce good aseptic results.
Given the economic importance of philodendrons and the need for sufficient micronutrients for growers of this plant, the aim of this study was to establish a protocol for establishing aseptic culture in vitro of philodendron xanadu to start mass reproduction and respond to the demand of this species.
Treatment of mother plants with sanitizers
Philodendron seedlings of 8 cm in height, obtained from in vitro culture and acclimatized, were established in 6-inch pots containing tezontle (red igneous rock of volcanic origin) with particle size of 5 mm as substrate. After transplanting, the plants were sprayed with sanitizing products. The plants were grown under greenhouse conditions. Sprays were applied every fourteen days for 11 months, together with fertilization; nutrition was applied in water according to Steiner’s formulation , and 1 mL L-1 of ANIBAC 580® (quaternary ammonium) was added to it to prevent the presence of pathogens in the substrate. Ten plants were used per treatment.
Murashige and Skog culture media with 30 g / l sucrose, 80 mg / l adenine sulfate, thiamine, glycine, pyridoxine, nicotinic acid (0.5 mg / l) and inositol (100 mg / l) were used. ; The pH was adjusted to 5.7 and 7.5 g / L Merck® agar was used as a gelling agent. Also, 100 ml flasks with 20 ml of culture medium were used. The culture medium was sterilized for 18 minutes in a vertical autoclave at 1.2 kg / cm2 at 120 ° C.
For in vitro establishment, plants treated with the sanitizers and grown for 185 days after transplanting were used. Axillary shoots of 3 to 5 cm were taken and placed separately according to treatment.Leaves and petioles were removed from each shoot and washed with plenty of soap and water; subsequently, the shoots were submerged for 30 min in a solution prepared with the fungicide Tokat® (1 mL L-1) and the bactericide Agry-Gent Plus 5000®. After 30 min, under aseptic conditions, the explants were immersed in 70% ethanol for 1 min, the alcohol was removed and a 0.16% sodium hypochlorite (NaOCl) solution was added; the explants in the solution were shaken for 5 min and then rinsed three times with sterile distilled water. in height; five explants were established per culture flask and incubated at 28 ± 2 ºC, with a photoperiod of 16 h light and 8 h dark, and a light intensity of 32 µmol·m 2·s-1.
Identification of contaminants
To identify the contaminants that grew during the establishment of the aseptic philodendron culture, a sample of each contaminated explant was taken with a sterile dissection needle; 12 fungal isolates and one bacterial isolate were obtained. Each sample was placed in a Petri dish with potato dextrose agar (PDA) and incubated at room temperature for 72 hours. The contaminants were reisolated again on PDA to obtain pure cultures. The fungal mycelium was mounted on slides with lactophenol blue. The morphological structures of the contaminant were observed with a compound microscope at 40x.
The pure isolate visually identified as bacterial was inoculated on PDA medium with 2 mL L-1 of 25% lactic acid and culture medium with eosin methylene blue (EMB) agar and incubated for 24 to 48 h. Isolations were also made in potato slices in humidity chambers using paper towels, water and sterile Petri dishes; a scratch was made with a sterilized dissection needle and inoculated with the pure culture.