Overexpression of 1-Aminocyclopropane-1-Carboxylic Acid Deaminase (acdS) Gene in Petunia hybrida Improves Tolerance to Abiotic Stresses

Abiotic stress induces the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) in plants, which consequently enhances ethylene production and inhibits plant growth. The bacterial ACC deaminase enzyme encoded by the acdS gene reduces stress-induced ethylene production and improves plant growth in response to stress. In this study, overexpression of acdS in Petunia hybrida (‘Mirage Rose’) significantly reduced expression of the ethylene biosynthesis gene ACC oxidase 1 (ACO1) and ethylene production relative to those in wild type (WT) under various abiotic stresses (cold, drought, and salt). The higher reduction of stress-induced ethylene in the transgenic plants, which was due to the overexpression of acdS, led to a greater tolerance to the stresses compared to that in the WT plants. The greater stress tolerances were proven based on better plant growth and physiological performance, which were linked to stress tolerance. Moreover, expression analysis of the genes involved in stress tolerance also supported the increased tolerance of transgenics relative to that with the WT. These results suggest the possibility that acdS is overexpressed in ornamental plants, particularly in bedding plants normally growing outside the environment, to overcome the deleterious effect of ethylene on plant growth under different abiotic stresses. The development of stress-tolerant plants will be helpful to advance the floricultural industry.

A study on the primary materials for in vitro propagation of Petunia hybrida L.

The aim of this study was to identify the primary materials for in vitro propagation of the Petunia hybrida L. The sterilisation method and the medium in different stages: shoot regeneration, rapid shoot multiplication, and the part of the in vitro plant used for rapid multiplication were identified. The study results showed that using nodal segments with axillary buds as the primary material and sterilising by HgCl2 0.1% in 10 minutes gave the best disinfection effect (survival rate reached 60%). Murashige and Skoog (MS) medium supplemented with 1 mg of BAP/l + 6.5 g agar/l + 30 g sacarose/l was reported as the best one for shoot regeneration (100% of shoot formed and number shoot per sample was 2.73), meanwhile, the mixture of MS + 1 mgBA/l + 0.2 mg IBA/l + 6.5 g agar/l + 30 g sacarose/l was considered as the best medium for rapid shoot multiplication (average shoots/sample reached 19.53). In vitro,leaf tissue is the most suitable vegetative organ for rapid multiplication, 100% of in vitro shoots regenerate through the form of callus, and the ratio of shoots/sample reached 21.53.

Cloning and Expression Analysis of Flavonoid 3′, 5′-Hydroxylase Gene from Brunfelsia acuminata

The full-length cDNA sequence of F3′5′H gene from the Brunfelsia acuminata was obtained by RT-PCR and RACE, whose GenBank accession number is JQ678765. The sequence contains a 1521 bp open reading frame, 120 bp 5′UTR and 61 bp 3′UTR, encoding a total of 506 amino acids. The molecular mass of the predicted protein is 56.47 kDa with an estimated pI of 8.78, respectively. Sequence alignment showed that the amino acid sequence of F3′5′H was 91%, 87% and 84% with that of Petunia × hybrida, Nierembergia sp., Solanum tuberosum, respectively. Real-time quantitative PCR analysis showed that the expression of F3′5′H gene was different in petals of different days, which was the highest expression level on day 0 and significantly higher than other days. The results indicated that F3′5′H might play key role in flower color regulation and provide a theoretical reference for blue flower molecular breeding.

Identification of WRKY transcription factors associated with leaf and corolla senescence in Petunia hybrida

Several families of transcription factors (TFs) control the progression of senescence. Many key TFs belonging to the WRKY family have been described to play crucial roles in the regulation of leaf senescence, mainly in Arabidopsis. However, little is known about senescence-associated WRKY members in floricultural species. Delay of senescence in leaves and petals of Petunia hybrida, a worldwide ornamental crop are highly appreciated traits. In this work, starting from 28 differentially expressed WRKY genes of Arabidopsis during the progression of leaf senescence, we identified the orthologous in P. hybrida and explored the expression profiles of 20 PhWRKY genes during the progression of natural (age-related) leaf and corolla senescence as well as in the corollas of flowers undergoing pollination-induced senescence. Simultaneous visualization showed consistent and similar expression profiles of PhWRKYs during natural leaf and corolla senescence, although weak expression changes were observed during pollination-induced senescence. Comparable expression trends between PhWRKYs and the corresponding genes of Arabidopsis were observed during leaf senescence, although more divergences were found in petals of pollinated petunia flowers. Integration of expression data with phylogenetics, conserved motif and cis-regulatory element analyses were used to establish a list of solid candidates that could regulate more than one senescence process. Our results suggest that several members of the WRKY family of TFs are tightly linked to the regulation of senescence in P. hybrida.

Effects of Maternal Environment on Seed Germination and Seedling Vigor of Petunia × hybrida under Different Abiotic Stresses

Seed germination and seedling vigor can be affected by environmental cues experienced by the mother plant. However, information about how the maternal environment affects seed quality is scarce in ornamental plants. This study aimed to investigate the effects of two different maternal environments on the seed germination and seedling vigor of Petunia × hybrida under a variety of abiotic stresses. Petunia mother plants were grown in either a greenhouse during the summer months or an indoor controlled-temperature-and-light environment. Collected seeds were subjected to external stressors, including polyethylene glycol (PEG), sodium chloride (NaCl), high temperature, and abscisic acid (ABA), to determine seed germination percentage and seedling vigor. Results indicated that seeds harvested from the mother plants grown in a controlled environment germinated better than seeds harvested from the mother plants grown in the greenhouse when suboptimal germination conditions were applied. Additionally, the seedlings from the controlled maternal environment performed better in both ABA and salinity stress tests than the greenhouse seedlings. Interestingly, the greenhouse seedlings displayed less reactive oxygen species (ROS) damage and lower electrolyte leakage than the controlled environment seedlings under dehydration stress. The difference in germination and seedling vigor of seeds from the two different maternal environments might be due to the epigenetic memory inherited from the mother plants. This study highlighted the strong impact of the maternal environment on seed germination and seedling vigor in Petunia and may assist in high-quality seed production in ornamental plants.

The Senescence-Associated Endonuclease, PhENDO1, Is Upregulated by Ethylene and Phosphorus Deficiency in Petunia

The upregulation of endonuclease activities and subsequent decreases in the nucleic acid content of leaves and petals are characteristics of senescence that allow for nutrient remobilization from dying organs. We previously identified a 43-kDa endonuclease activity (PhNUC1) that was upregulated in Petunia × hybrida petals during senescence. PhNUC1 has optimal activity at neutral pH, is enhanced by Co2+, and degrades both DNA and RNA. The peptide sequence of a 43-kDa endonuclease identified from senescing petals by 2-dimensional gel electrophoresis was used to clone the gene (PhENDO1) encoding the senescence-associated protein. PhENDO1 expression was upregulated in petals during the senescence of unpollinated and pollinated flowers and by ethylene treatment. When petunias were grown under nutrient deficient conditions, P-starvation, and to a lesser extent N-starvation, induced expression of PhENDO1. The endogenous expression of PhENDO1 was down regulated using virus induced gene silencing (VIGS), and in-gel endonuclease assays confirmed that the activity of the 43-kDa PhNUC1 was decreased in senescing corollas from PhENDO1-silenced (pTRV2:PhCHS:PhENDO1) plants compared to controls (pTRV2:PhCHS). Down regulating PhENDO1 in petunias did not alter flower longevity. While PhENDO1 may be involved in nucleic acid catabolism during senescence, down regulating this gene using VIGS was not sufficient to delay flower senescence.