Abstract
BackgroundCymbidium faberi is one of the oldest cultivars of oriental orchids, with an elegant flower fragrance. In order to investigate the molecular mechanism and the functions of related proteins in the methyl jasmonate (MeJA) signaling pathway, one of the main components of flower fragrance in C. faberi, yeast one- and two-hybrid three-frame cDNA libraries were constructed.ResultsIn this study, a modified cDNA library used for yeast one- and two-hybrid screening was successfully constructed, with a recombinant efficiency of 95%. The lengths of inserted fragments ranged from 750~3000 bp, and the library capacity reached 6 × 109 CFU/ μg of cDNA insert, which was suitable for the requirements of subsequent screening. Finally, a homologous protein related with pathogenesis was screened out by the bait vector of CfbHLH36, which may participate in the MeJA signaling pathway.ConclusionThe yeast one- and two-hybrid library of C. faberi provides large amounts of useful information for the functional genomics research in C. faberi, and this method could also be applied to other plants to screen DNA-protein and protein-protein interactions.
Highlights
Cymbidium faberi is one of the oldest cultivars of oriental orchids, with an elegant flower fragrance
Cymbidium faberi is famous for its soft color and strong flower fragrance
The mRNA was reverse-transcribed into first-strand cDNA, which was used to synthesize the double-stranded cDNA by Long distance-PCR (LD-PCR) (Fig. 2b)
Summary
Cymbidium faberi is one of the oldest cultivars of oriental orchids, with an elegant flower fragrance. In order to investigate the molecular mechanism and the functions of related proteins in the methyl jasmonate (MeJA) signaling pathway, one of the main components of flower fragrance in C. faberi, yeast one- and two-hybrid three-frame cDNA libraries were constructed. Orchids are generally divided into tropical and oriental cultivars. Cymbidium faberi is famous for its soft color and strong flower fragrance. The wild populations have significantly deteriorated due to over-exploitation. In order to develop new cultivars of Cymbidium via genetic engineering and preserve the wild resources, it is imperative to elucidate the biosynthetic pathways and molecular mechanisms of its economically important traits. The yeast one- and two-hybrid systems are commonly used to screen for interactions between target proteins
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