Effect Of Anthocyanin Accumulation In Nutrient Organs On The Ability Of Petunia Hybrida To Tolerate Low Temperature And UV-B Radiation

Petunia hybrida is an annual or perennial herb in the genus Pittosporum,and a common indoor flowering and outdoor greening plant with a wide range of applications.The improvement of ornamental traits of Petunia mainly focuses on flower color and plant shape,and less attention is paid to the improvement of leaf color.But with the continuous development of urban garden construction,the ornamental characteristics of Petunia can no longer meet the needs of people.The accumulation of anthocyanin,a water-soluble pigment present in plant cytosol,affects the coloration of petals,fruits and leaves,and some studies have also shown that anthocyanin is closely related to plant stress resistance.Therefore,in this paper,we investigated the correlation between Petunia leaf color improvement and anthocyanin accumulation and stress resistance based on anthocyanin anabolic pathway,and the results are as follows:(1)Application of Arabidopsis green tissue-specific promoter-driven ANTHOCYANIN 4(AN4)gene expression in Petunia leaf with color improvement: The green tissue-specific promoter of CHLOROPHYLL A/B-BINDING PROTEIN 3(CAB3)was amplified from the Arabidopsis,and a plant expression vector p G536-CAB3 was constructed for this promoter and AN4,a transcription factor related to Petunia anthocyanin synthesis.CAB3:AN4 was transferred into Petunia under the mediation of Agrobacterium tumefaciens,and seven independent genetic strain were obtained.The phenotypic changes of the transgenic plants were not obvious,only the young leaves were slightly purple.(2)Application of-estradiol chemically induced expression vector system in Petunia:The study introduced XVE(Lex A-VP16-ER),an estrogen-based system for inducing the expression of target genes,and constructed the vector p ER8:AN4 that regulates the induction of AN4 gene expression,which was introduced into Petunia using the Agrobacterium-mediated leaf disc method.Positive transformed strains were identified by hygromycin screening and PCR,then obsered the different of the positive transformed strains by applying different concentrations of β-estradiol to different organs.For the roots,it was found that 2 μM could induce the expression of AN4 gene with a significant effect and the induction efficiency was75%.For the leaf,the induction efficiency was 88.2% with 20 μM,and the observation time of10 d was accurate.The best effect and the longest duration were achieved by applying 20 μM estradiol after 10 d of preculture of leaf,and the most obvious change in leaf color was observed around the 4th d after induction,and it is presumed that the maximum expression of exogenous gene induction was reached at this time.(3)Effect of anthocyanin accumulation in nutritional organs on low temperature tolerance of Petunia: Five transgenic Petunia were used for the study,and the Chlorophyll a,Chlorophyll b,Carotenoids,Chlorophyll,Relative conductivity,Malondialdehyde,Soluble protein,Soluble sugar,Superoxide dismutase,Peroxisomal,and Peroxidase were measured between the purple leaves and other green leaf strains by natural low temperature treatment at the yong stage of Petunia.The 10 physiological indexes were divided into three principal component factors(with a cumulative contribution of 88.659%)using principal component analysis,and the score of each transgenic material was calculated as AN40E-1> AN40E-3>an4>0229-15>dpl by combining the fuzzy affiliation function method.The results showed that the adaptability of purple-leaved transgenic Petunia was stronger than that of green-leaved Petunia under natural low temperature conditions,and anthocyanin accumulation may improve the environmental adaptability of Petunia.(4)Effect of anthocyanin accumulation in nutrient organs on Ultraviolet radiation-B(UV-B)tolerance in Petunia: Seven transgenic Petunia(p ER8:AN4 was divided into foliar β-estradiol treatment group A and control group B)were used under UV-B radiation.Four physiological indicators of resistance,including Chlorophyll,Carotenoids,Relative conductivity,and Relative water content,were measured in young leaves,and to evaluate the tolerance to UV-B.The score was CAB3:AN4>AN4OE>an4>p ER8:AN4-A>dpl>0229-15>p ER8:AN4-B.The results showed that a short period of foliar spraying with 20 μM could significantly increase the resistance of the p ER8:AN4 strain to UV-B.The accumulation of anthocyanin in nutritional organs was consistent with the ability of Petunia to tolerate UV-B,the excessive accumulation of anthocyanin in leaves may affect the growth.