Selection and validation of appropriate reference genes and expression analysis of jasmonic acid-related genes responding to Marssonina rosae in Rosa species and cultivars

  Objective  This paper aims to select the appropriate reference gene and analyze the role of jasmonic acid in Rosa species and cultivars responding to Marssonina rosae.

  Method  R. multiflora f. carnea, R. xanthina f. spontanea, R. glauca, R. rugosa, R. hybrida ‘Porcelina’ and R12-26 (hybrid of R. rugosa and R. hybrida ‘Porcelina’) were used as materials, qRT-PCR technology, geNorm, NormFinder, BestKeeper were used to evaluate the expression stability of nine candidate reference genes (ACT, GAPDH, PP2A, RCl2, SAND, TIP, TUA, TUB and UBC). The expression levels of jasmonic acid resistance pathway relatedgenes (COI1, OPR3, MYC2 and JAR1) in six Rosa species and cultivars were quantitatively analyzed.

  Result  (1) UBC was the common reference gene of six Rosa species and cultivars responding to M. rosae, and can be used for analyzing the related gene expression levels to JA resistance pathway. (2) There were differences in JA content among the six plants responding to M. rosae. In disease-resistant plants, JA concentration decreased at 0−4 d and increased at 4−8 d. In disease-susceptible plants, JA concentration decreased at 0−8 d and increased significantly at 8−10 d. (3) The expression levels of JA synthesis-related genes OPR3 and JAR1 were different at the early stage of infection. The expression level of OPR3 was down-regulated and JAR1 up-regulated in five plants except for R. multiflora f. carnea. OPR3 and JAR1 were all up-regulated in the late stage of infection, however the up-regulation degree in black-spot susceptible species was significantly higher than that of resistant species. (4) There were also differences in the expression levels of JA signaling related genes COI1 and MYC2 at the early stage of infection. The expression of COI1 was up-regulated in disease-resistant plants and down-regulated in disease-susceptible plants at the early stage of infection, MYC2 was down-regulated in 6 plants infected 0−2 d. COI1 and MYC2 were up-regulated after 2 d of infection, and the up-regulation degree in black-spot susceptible species was significantly higher than that of resistant species.

  Conclusion  MYC2 and COI1, which are related to JA signaling, play a negative regulatory role in the resistance of Rosa plants to the invasion of black spot pathogens, and the JA pathway mediated resistance to the invasion of dead vegetative pathogens plays a role in the later stage of infection.