Random sequencing of sweet orange (Citrus sinensis Osbeck) cDNA library derived-from young seeds

Abstract

Sequencing of randomly selected cDNA clones was carried out to survey the repertoire of genes expressed in immature seeds of 'Valencia' orange (Citrus sinensis Osbeck) and to produce restriciton fragment length polymorphism (RFLP) markers on the genome mapping program in Citrus. A cDNA library was made by using mRNA prepared from immature seeds of 'Valencia' orange whose developing embryos were at an early globular state. A total of 192 individual cDNA clones were partially sequenced and compared with the GenBank database. The cDNA clones, which presented significant homology to previously characterized plant, eukaryote, and all other organism genes (optimized (OPT) score >200) in data source formed 24.5% (47 clones), 12.0% (23 clones) and 8.8% (17 clones) of total cDNA clones analyzed in this experiment. The seed cDNA library showed low level of redundancy of cDNA clones. Some clones such as metallothionein, however, were revealed several times, suggesting possible isoforms. Although putative genes, which are though to play a role on embryogenesis, were not screened in this experiment, a wide range of genes involved in functions such as metabolism, translation, signal transduction, and stress responses were obtained. The randomly sequenced cDNA clones described in this study provide, for the first time, and indication of the types of genes expressed at an early stage of seed development in Citrus. Furthermore, this method made possible the selection of many gene species which could be used as RFLP probes.