Holistic protocol for callus culture optimization using statistical modelling

Abstract

Plants endue a key role against illnesses caused by oxidative stress. These attributes are frequently associated with polyphenolic compounds. However, presence and concentration of secondary metabolites are affected by abiotic factors. The in vitro culture techniques can solve these drawbacks. Peppers can be a suitable alternative to obtain polyphenols. Aiming to optimise the callus culture stage from Capsicum baccatum to produce polyphenols, this work evaluated systemically the effects of the explant's origin (root, hypocotyl and cotyledon), growth hormone type (2,4-dichlorophenoxyacetic acid (2,4-D), benzylaminopurine (BAP) and a combination of 2,4-D/BAP at five-to-one ratio) and concentration (0.023-10.000 mg L) on callus culture efficiency parameters using a multilevel factorial design. The root explant in combination with BAP at 1.138 mg L ensured the optimal values of the assessed responses; ​callus mass (225.03 mg), antioxidant activity (35.95%), total phenols (11.48 mg of GAE/g DE) and flavonoids (15.92 mg of RU/g DE) production.

Handling culture medium composition for optimizing plant cell suspension culture in shake flasks

“…1b) within the experimental domain under consideration. Besides, the antioxidant activities herein (suspension cell culture) were lower than those previously obtained for our group in callus culture with homologous basal culture medium and hormone supplementation, as well as, explant origin (7.58 ± 4.07%) (Fidemann et al 2017).…”

“…The detailed procedures for the culture initiation were previously reported (Fidemann et al 2016). With the aid of sterilized tweezers and scissors, explants were cut and inoculated in Petri dishes (15 × 100 mm) (five explants per dish) containing 35 mL Murashige and Skoog culture medium (MS) (pH 5.75) (Murashige and Skoog 1962) solidified with Phytagel (Sigma Chemical Co., St. Louis, MO) 4 g L −1 , supplemented with 3% sucrose and 2,4-d/BAP (5:1 ratio) at 1.138 mg L −1 (these conditions were optimized in previous work of our team) (Fidemann et al 2017). The culture media were previously autoclaved at 121 °C, 1 atm for 20 min.…”

“…However, equivalent studies including C. baccatum species are scarce. Our group has been recently focused on the initial in vitro culture steps (in vitro germination and callus culture) for obtaining antioxidant compounds from C. baccatum species (Fidemann et al 2016(Fidemann et al , 2017.…”