Abstract
BackgroundAsian Palmyra palm, the source of palm-sugar, is dioecious with a long juvenile period requiring at least 12 years to reach its maturity. To date, there is no reliable molecular marker for identifying sexes before the first bloom, limiting crop designs and utilization. We aimed to identify sex-linked markers for this palm using PCR-based DNA fingerprinting, suppression subtractive hybridization (SSH) and transcriptome sequencing.MethodsDNA fingerprints were generated between males and females based on RAPD, AFLP, SCoT, modified SCoT, ILP, and SSR techniques. Large-scale cloning and screening of SSH libraries and de novo transcriptome sequencing of male and female cDNA from inflorescences were performed to identify sex-specific genes for developing sex-linked markers.ResultsThrough extensive screening and re-testing of the DNA fingerprints (up to 1,204 primer pairs) and transcripts from SSH (>10,000 clones) and transcriptome data, however, no sex-linked marker was identified. Although de novo transcriptome sequencing of male and female inflorescences provided ∼32 million reads and 187,083 assembled transcripts, PCR analysis of selected sex-highly represented transcripts did not yield any sex-linked marker. This result may suggest the complexity and small sex-determining region of the Asian Palmyra palm. To this end, we provide the first global transcripts of male and female inflorescences of Asian Palmyra palm. Interestingly, sequence annotation revealed a large proportion of transcripts related to sucrose metabolism, which corresponds to the sucrose-rich sap produced in the inflorescences, and these transcripts will be useful for further understanding of sucrose production in sugar crop plants. Provided lists of sex-specific and differential-expressed transcripts would be beneficial to the further study of sexual development and sex-linked markers in palms and related species.
Highlights
Asian Palmyra palm (Borassus flabellifer L, 2n = 36) is a dioecious and slow-growing perennial tree, requiring 12–15 years to reach its maturity and produce the first inflorescence (Kovoor, 1983)
George et al (2007) had developed a RAPD based male-specific marker (OPA06600) for the Asian Palmyra palm population in India. We tested this marker for sex identification in our population in Thailand using up to 20 male and 24 female individuals, but this marker was unable to confirm the sexes (Fig. S1)
Despite such extensive screening of DNA markers up to 1,204 primer pairs by the ten techniques, we did not obtain any sex-linked marker. This experiment showed that the DNA fingerprinting covered here is inadequate for identifying a sex-linked marker for Asian Palmyra palm
Summary
This palm is found widespread in South and Southeast Asia and provides essential food and economic values throughout its parts including inflorescence sap for producing palm sugar and alcoholic beverages, fruits for consumption and the tree trunk for construction (Morton, 1988; Lim, 2012) Both sexes are morphologically identical except for the male and female inflorescences, and there are no reliable means of sex identification before the first bloom. De novo transcriptome sequencing of male and female inflorescences provided ∼32 million reads and 187,083 assembled transcripts, PCR analysis of selected sex-highly represented transcripts did not yield any sex-linked marker This result may suggest the complexity and small sexdetermining region of the Asian Palmyra palm. Provided lists of sex-specific and differential-expressed transcripts would be beneficial to the further study of sexual development and sex-linked markers in palms and related species
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