Differential expression of Isochorismate synthase in Catharanthus roseus during ‘Candidatus Liberibacter asiaticus’ infection

Abstract

Periwinkle facilitates study of the non-culturable pathogen 'Candidatus Liberibacter asiaticus' (CaLas), the causal agent of Huanglongbing (HLB) in citrus. Isochorismate synthase (ICS) is involved in the defense responses to pathogens through the salicylic acid (SA) signaling pathway in some plants. In this study, crICS from Catharanthus roseus was identified. The fulllength cDNA of crICS is 2206 bp long, with an open reading frame (ORF) encoding a putative protein of 322 amino acids, including a conserved chorismate binding enzyme motif. Phylogenetic analysis revealed the crICS protein to cluster together with those of Rubia cordifolia and Coffea canephora to form a separate clade. Results of a qRT-PCR analysis indicated that crICS was constitutively expressed in leaf, stem, and root tissue of healthy periwinkle plants. During CaLas infection, crICS expression levels increased gradually and reached a peak at 22 days after inoculation (DAI), and then decreased gradually but were still higher than the expression level for the control group (CK) until the end of the experiment. To investigate whether crICS may act to mediate resistance through the SA signaling pathway in periwinkle, we sprayed SA on leaves, and found that SA increased crICS expression levels, but did not decrease the cumulative CaLas-positive rate of infected plants. The results suggest that crICS most likely mediates resistance to CaLas through the SA signaling pathway.

Expression of PEI-coated gold nanoparticles carrying exogenous gene in periwinkle mesophyll cells and its practice in huanglongbing research

“…The sequences of the primers used were collated in Table S1 . The qPCR conditions were as follows: 94°C for 2 min, followed by 30 cycles of 94°C for 20 s, 60°C for 20 s, and 72°C for 20 s. The qPCR data were analyzed by the ddCt method to obtain the normalized gene expression fold change with respect to the 18S rDNA housekeeping gene and control sample ( Li et al., 2019c ; Demirer et al., 2019 ). For each sample, qPCR was performed as three technical replicates (three reactions from the same isolated RNA batch), and the entire experiment consisting of independent infiltrations and RNA extractions from different plants was repeated three times (three biological replicates).…”

Changes in the expression of CrFTA, the Catharanthus roseus farnesyltransferase α-subunit gene, in response to a Candidatus Liberibacter asiaticus infection

“…The total RNA was extracted from the samples using a Trizol Reagent (Invitrogen, CA, USA). The genomic DNA was extracted from 0.1 g of the leaf tissue sampled from the same plants at the same time using the method of Li et al (2019).…”