In vitro cluster buds regeneration and control of shoot tip necrosis in tissue cultures of Trichosanthes cucumerina L.

Trichosanthes cucumerina L. is a medicinal vine with great potential that can be used as a vegetable. In this study, an efficient in vitro regeneration system of T. cucumerina was established by inducing adventitious buds and controlling Shoot Tip Necrosis (STN) in the in vitro culture. The results of hormone single factor and orthogonal experiments showed that 6-BA had a significant effect on inducing adventitious buds. The best medium for bud proliferation was MS + 6-BA 3.0 mg·L− 1 + KIN 2.0 mg·L− 1 + NAA 0.5 mg·L− 1, and the proliferation capacity was 14.68. However, after three consecutive subcultures, a higher incidence of STN (78.86%) was observed. The results of the STN control experiment showed that reducing the concentration of inorganic salt, shortening the subculture interval, and increasing the Ca2+ in the medium could effectively control the occurrence of STN. However, reducing the concentration of inorganic salt and shortening the subculture interval excessively reduces the proliferation capacity significantly. Considering both the incidence of STN and proliferation capacity, the measures proposed in this study to alleviate STN were to keep the concentration of inorganic salt unchanged, adjust the subculture interval from 45 days to 40 days, and increase the Ca2+ concentration in the medium to 480 mg·L− 1. When the subculture interval was 40 d, the stem segments obtained the lowest STN incidence (8.3%) and higher shoot proliferation capacity (10.02) on the medium of MS + 3.0 mg·L− 1 6-BA + 2.0 mg·L− 1 KIN + 0.5 mg·L− 1 NAA + 360.36 mg·L− 1 Ca2+ (360 mg·L− 1 is the concentration of additional calcium to be added, and after this addition, the total calcium concentration in the MS medium was 480.48 mg·L− 1). The best rooting medium for stem segments was MS + 0.50 mg·L− 1 NAA; the rooting rate was 100%, and no STN occurred. After acclimation, the survival rate of the rooted plantlet reached 95% after 40 days in the soil matrix of coconut husk: humus soil: perlite = 2: 2: 1 (v/v). The results of this study not only reveal the mechanism of STN in T. cucumerina, but also provide technical support and a theoretical basis for its further breeding and cultivation. In this study, an efficient and stable tissue culture rapid propagation system for Trichosanthes cucumerina L. was formed by controlling the shoot tip necrosis in vitro culture.