An Examination of Differentially-Expressed Genes from Oil Palm Embryogenic and Non-Embryogenic Cultures

To date the embryogenesis rate in oil palm tissue culture averages 6%. Thus, there is a need to find ways to increase this rate and also to create a selection system that is able to distinguish embryogenically competent calli from their non-embryogenic counterpart at an early stage. Using cold plaque screening of oil palm suspension cultures, about 1000 clones were isolated. About sixty-four percent of the clones have extremely low expression levels in suspension cultures and non-embryogenic calli. Another 22% were found to be up-regulated in suspension cultures compared to non-embryogenic calli. Out of the 600 sequenced clones, 46% were found to be novel or similar to proteins with unknown functions. Most of the other genes were found to be involved in cell metabolism and proliferation, which leads to an increase in the expression of genes involved in namely, protein synthesis and signal transduction pathways. cp919.1, a TUBBY-like protein homolog or a similar member of the TULP family is up-regulated in non-embryogenic callus. cp 194.2, a member of a novel serine/threonine kinase subfamily, is up-regulated in embryogenic callus. As hypothesized in the mammalian systems, cp194.2 may play a role in the disruption of the extracellular matrix surrounding the proembryogenic masses. Transcripts of cp664.2, a truncated leucine-rich-repeat encoding protein and cp610.2, a HD-Zip II gene were up-regulated in suspension cultures. The role of cp610.2 is unknown, but it may be involved in the transcription of genes involved in early embryogenesis. 3N42.2 was isolated by RT-PCR followed by cDNA library screening. 3N42.2, a NAC superfamily member, like CUCI/2 may be expressed in the presumptive shoot apical meristem of the embryos and then be restricted to the boundary regions of the apical meristem at later embryogenic stages. 3N42.2 expression then remains at the boundary regions of the meristem during post-embryonic development. cp194.2, cp610.2 and 3N42.2 may be used as markers for early somatic embryogenesis. Functional elucidation needs to be carried out to understand the roles that these proteins play in plant development

Overexpression of the oil palm (Elaeis guineensis Jacq.) TAPETUM DEVELOPMENT1-like Eg707 in rice affects cell division and differentiation and reduces fertility

The functional analysis of the TAPETUM DEVELOPMENT1-like analog Eg707 of oil palm was carried out in rice by over-expressing Eg707 under the control of a double cauliflower mosaic virus 35S promoter. Ectopic expression of Eg707 in rice induced dark green and matured compact brownish calli compared to pale wild type and negative control calli. Regenerated transgenic rice plants exhibited a reduction in organ size and plant height, rolled, erect leaves, less tillers, increased chlorophyll content, and reduced fertility with smaller green seeds. At the molecular level Eg707 overexpression caused an increase in the transcription of SAPK9, a SnRK2 protein kinase family member that is activated by ABA and hyperosmotic stress. Together, the results show that ectopic Eg707 expression influences cell division and differentiation, presumably via altered hormone homeostasis

A model for predicting flower development in Elaeis guineensis Jacq

The proper development of oil palm fruit is important as the source of oil is the fruit mesocarp and kernel. Prior to fruit formation, the development of flowers is therefore also important. Determination of the flower development stages in oil palm generally involves tedious histological analyses of each sampled inflorescence, making it a costly and inefficient way of gauging the developmental state. In this study, a statistical model was established from the association of physical or macroscopic measurement data to flower development, which was determined via histological analyses. The final reduced ordinal logistic regression model is a partial proportional odds model that uses inflorescence length and palm age as predictors to predict the flower development stage. The likelihood-ratio χ2 test suggested the model adequately fits the data (p<0.01). The model, with a prediction accuracy of 78.5%, can be used for selecting inflorescences of specific development stages from palms aged three to 10 years of field-planting. These stages can be further verified by histological analyses. This lowers the overall costs and time by reducing the number of samples requiring histological analysis prior to downstream studies

A novel transcript of oil palm (Elaeis guineensis Jacq.), Eg707, is specifically upregulated in tissues related to totipotency

In this study, we report the molecular characterization of clone Eg707 isolated from cell suspension culture of the oil palm. The deduced polypeptide of clone Eg707 is highly similar to an unknown protein from Arabidopsis thaliana. The presence of an Ald-Xan-dh-C2 superfamily domain in the deduced protein sequence suggested that Eg707 protein might be involved in abscisic acid biosynthesis. Eg707 might be present as a single copy gene in the oil palm genome. This gene is highly expressed in tissue cultured materials compared to vegetative and reproductive tissues, suggesting a role of this gene during oil palm somatic embryogenesis or at the early stages of embryo development. Expression analysis of Eg707 by RNA in situ hybridization showed that Eg707 transcripts were present throughout somatic embryo development starting from proembryo formation at the embryogenic callus stages till the maturing embryo stages. Since proembryo formation within the embryogenic callus is one of the first key factors in oil palm somatic embryo development, it is suggested that Eg707 could be used as a reliable molecular marker for detecting early stage of oil palm somatic embryogenesis

RNA from fresh frozen cryosections of oil palm inflorescences is superior to FFPE sections

Formalin-fixed, paraffin-embedded (FFPE) and fresh frozen (FF) tissue sections are valuable sources for histology and molecular studies. However, FFPE tissue sections usually provide poor quality RNA that is unsuitable for most downstream molecular applications. In this study, tissue preparation for cryosectioning and RNA extraction protocols were optimised for FF oil palm inflorescences. FF tissue treated with sucrose offered good cellular image resolution and more importantly, good RNA quality. Higher RNA quality with good RIN values (>6) was obtained from FF sections compared to the fragmented RNA from FFPE tissue sections. However, FFPE sections provided better resolution in terms of cellular morphology. The quality of cellular morphology of the FF sections was moderate, but was sufficient to distinguish the different cell types. Hence, FF oil palm inflorescences is a better choice for downstream transcriptomics studies that require tissue sections without compromising the cytological details

Innate Immune Responses of Pulmonary Epithelial Cells to Burkholderia pseudomallei Infection

10.1371/journal.pone.0007308PLoS ONE410

The Core and Accessory Genomes of Burkholderia pseudomallei: Implications for Human Melioidosis

Natural isolates of Burkholderia pseudomallei (Bp), the causative agent of melioidosis, can exhibit significant ecological flexibility that is likely reflective of a dynamic genome. Using whole-genome Bp microarrays, we examined patterns of gene presence and absence across 94 South East Asian strains isolated from a variety of clinical, environmental, or animal sources. 86% of the Bp K96243 reference genome was common to all the strains representing the Bp “core genome”, comprising genes largely involved in essential functions (eg amino acid metabolism, protein translation). In contrast, 14% of the K96243 genome was variably present across the isolates. This Bp accessory genome encompassed multiple genomic islands (GIs), paralogous genes, and insertions/deletions, including three distinct lipopolysaccharide (LPS)-related gene clusters. Strikingly, strains recovered from cases of human melioidosis clustered on a tree based on accessory gene content, and were significantly more likely to harbor certain GIs compared to animal and environmental isolates. Consistent with the inference that the GIs may contribute to pathogenesis, experimental mutation of BPSS2053, a GI gene, reduced microbial adherence to human epithelial cells. Our results suggest that the Bp accessory genome is likely to play an important role in microbial adaptation and virulence

Molecular characterization of an unknown protein (Acc. No. EU795363) from the ESTs of oil palm (Elaeis guineensis Jacq.) cell suspension culture.

A large quantity of ESTs is available from various cDNA libraries of oil palm. The information from oil palm EST databases has been utilized to identify several interesting sequences for molecular characterization. In this study, we report molecular characterization of clone 583 (Acc. No. EU795363) isolated from cell suspension culture of oil palm. This clone is predicted to encode a single major open reading frame for a polypeptide of 177 amino acids with a predicted molecular mass of 19.6 kDa. The predicted amino acid sequence does not contain any signal peptide and transmembrane region. Based on Kyte-Doolittle hydropathy profile, this protein is predicted to be a soluble protein. The predicted ORF of clone 583 was 59% identical to an amino acid sequence of an unknown protein from Oryza sativa (Acc. BAD25663). Southern analysis showed that this clone might be a member of a multigene family in the oil palm genome. Gene expression study by real time quantitative RT-PCR showed that transcripts of clone 583 might be present in low abundance

Sequence and expression analysis of EgSAPK, a putative member of the serine/threonine protein kinases in oil palm (Elaeis guineensis Jacq.)

In present study, EgSAPK (EU805512), an oil palm transcript coding for a putative SAPK protein kinase, have been molecular characterized. The cDNA for EgSAPK isolated from an oil palm cell suspension culture is 1470 bp in length with a longest Open Reading Frame (ORF) of 963 bp. No translation start codon could be identified so EgSAPK cDNA sequence is lacking the 5’-end. The deduced protein sequence shares 89% identity with the serine/threonine protein kinase SAPK9 from rice (AB125310.1). Real time PCR results showed that the expression levels of EgSAPK varied in different oil palm tissues and the EgSAPK gene shares a similar expression pattern with the SAPK gene of rice. Furthermore, the transcription of the EgSAPK gene in green embryo, white embryo and embryogenic calli tissues were higher than in non-embryogenic calli tissues. Southern blot analysis showed that the EgSAPK gene might be present as a single copy gene in the oil palm genome. These results suggest that EgSAPK may have a similar function as the SAPK gene of rice and thus can be a candidate marker for oil palm somatic embryogenesis