Analysis of the Petunia hybrida double flower transcriptome using suppression subtractive hybridization

The aim of this study was to characterize the transcriptional response that is associated with the Petunia double flower phenotype with a view to understanding the underlying molecular mechanisms. We created forward and reverse subtracted cDNA libraries to represent the up-regulated genes found in the double and single flowers obtained from segregated progenies through backcross, respectively. This was achieved using suppression subtractive hybridization (SSH) followed by differential screening through dot blot hybridization. In total, 285 expressed sequence tags (ESTs) were identified. Analysis using Blastx searches showed that the libraries contained several important developmental genes. The expression patterns of selected genes, believed to be involved in the development of Petunia double flowers, were further analyzed by semi-quantitative or real-time reverse transcription-polymerase chain reaction (RT-PCR). These analyses were conducted using RNA extracts taken from specific floral organs and from flowers at various stages of development. Each of the selected genes showed differential expression levels between single and double flowers, and this was particularly clear in young flower buds, thereby suggesting that the genes are important for the early development of the double flower form. In particular, we discuss the expression patterns of genes with putative roles relating to cell expansion, organ fusion and organ number. Other genes, with less well-documented roles, may be involved in modulating the development of the Petunia double flower. Thus, such genes appear to function along with the major regulatory MADS-box genes as part of a complicated network of genetic factors that direct the formation of the Petunia double flower.

Research highlights▶ TM6, the B-class MADS-box member, was up-regulated in double Petunia. ▶ The up-regulation of PhFDH might contribute to the formation of double flower. ▶ In addition to the major regulatory MADS-box genes, the expression of other less well-documented genes may also be important to the complicated network of genetic factors directing the formation of the Petunia double flower.