Bioenergy

Wang, W, Messing J.  2015.  Status of duckweed genomics and transcriptomics. Plant Biol (Stuttg). 17 Suppl 1:10-5. AbstractWebsite
Duckweeds belong to the smallest flowering plants that undergo fast vegetative growth in an aquatic environment. They are commonly used in wastewater treatment and animal feed. Whereas duckweeds have been studied at the biochemical level, their reduced morphology and wide environmental adaption had not been subjected to molecular analysis until recently. Here, we review the progress that has been made in using a DNA barcode system and the sequences of chloroplast and mitochondrial genomes to identify duckweed species at the species or population level. We also review analysis of the nuclear genome sequence of Spirodela that provides new insights into fundamental biological questions. Indeed, reduced gene families and missing genes are consistent with its compact morphogenesis, aquatic floating and suppression of juvenile-to-adult transition. Furthermore, deep RNA sequencing of Spirodela at the onset of dormancy and Landoltia in exposure of nutrient deficiency illustrate the molecular network for environmental adaption and stress response, constituting major progress towards a post-genome sequencing phase, where further functional genomic details can be explored. Rapid advances in sequencing technologies could continue to promote a proliferation of genome sequences for additional ecotypes as well as for other duckweed species.
Wang, W, Wu Y, Messing J.  2014.  RNA-Seq transcriptome analysis of Spirodela dormancy without reproduction. BMC Genomics. 15:60. AbstractWebsite
BACKGROUND: Higher plants exhibit a remarkable phenotypic plasticity to adapt to adverse environmental changes. The Greater Duckweed Spirodela, as an aquatic plant, presents exceptional tolerance to cold winters through its dormant structure of turions in place of seeds. Abundant starch in turions permits them to sink and escape the freezing surface of waters. Due to their clonal propagation, they are the fastest growing biomass on earth, providing yet an untapped source for industrial applications. RESULTS: We used next generation sequencing technology to examine the transcriptome of turion development triggered by exogenous ABA. A total of 208 genes showed more than a 4-fold increase compared with 154 down-regulated genes in developing turions. The analysis of up-regulated differential expressed genes in response to dormancy exposed an enriched interplay among various pathways: signal transduction, seed dehydration, carbohydrate and secondary metabolism, and senescence. On the other side, the genes responsible for rapid growth and biomass accumulation through DNA assembly, protein synthesis and carbon fixation are repressed. Noticeably, three members of late embryogenesis abundant protein family are exclusively expressed during turion formation. High expression level of key genes in starch synthesis are APS1, APL3 and GBSSI, which could artificially be reduced for re-directing carbon flow from photosynthesis to create a higher energy biomass. CONCLUSIONS: The identification and functional annotation of differentially expressed genes open a major step towards understanding the molecular network underlying vegetative frond dormancy. Moreover, genes have been identified that could be engineered in duckweeds for practical applications easing agricultural production of food crops.
Calviño, M, Messing J.  2012.  Sweet sorghum as a model system for bioenergy crops.. Current opinion in biotechnology. 23(3):323-9. AbstractWebsite
Bioenergy is the reduction of carbon via photosynthesis. Currently, this energy is harvested as liquid fuel through fermentation. A major concern, however, is input cost, in particular use of excess water and nitrogen, derived from an energy-negative process, the Haber-Bosch method. Furthermore, the shortage of arable land creates competition between uses for food and fuel, resulting in increased living expenses. This review seeks to summarize recent knowledge in genetics, genomics, and gene expression of a rising model species for bioenergy applications, sorghum. Its diploid genome has been sequenced, it has favorable low-input cost traits, and genetic crosses between different cultivars can be used to study allelic variations of genes involved in stem sugar metabolism and incremental biomass.
Liao, JC, Messing J.  2012.  Energy biotechnology. Current opinion in biotechnology. 23(3):287-9.Website
Wang, W, Messing J.  2012.  Analysis of ADP-glucose pyrophosphorylase expression during turion formation induced by abscisic acid in Spirodela polyrhiza (greater duckweed). BMC Plant Biol. 12:5. AbstractWebsite
BACKGROUND: Aquatic plants differ in their development from terrestrial plants in their morphology and physiology, but little is known about the molecular basis of the major phases of their life cycle. Interestingly, in place of seeds of terrestrial plants their dormant phase is represented by turions, which circumvents sexual reproduction. However, like seeds turions provide energy storage for starting the next growing season. RESULTS: To begin a characterization of the transition from the growth to the dormant phase we used abscisic acid (ABA), a plant hormone, to induce controlled turion formation in Spirodela polyrhiza and investigated their differentiation from fronds, representing their growth phase, into turions with respect to morphological, ultra-structural characteristics, and starch content. Turions were rich in anthocyanin pigmentation and had a density that submerged them to the bottom of liquid medium. Transmission electron microscopy (TEM) of turions showed in comparison to fronds shrunken vacuoles, smaller intercellular space, and abundant starch granules surrounded by thylakoid membranes. Turions accumulated more than 60% starch in dry mass after two weeks of ABA treatment. To further understand the mechanism of the developmental switch from fronds to turions, we cloned and sequenced the genes of three large-subunit ADP-glucose pyrophosphorylases (APLs). All three putative protein and exon sequences were conserved, but the corresponding genomic sequences were extremely variable mainly due to the invasion of miniature inverted-repeat transposable elements (MITEs) into introns. A molecular three-dimensional model of the SpAPLs was consistent with their regulatory mechanism in the interaction with the substrate (ATP) and allosteric activator (3-PGA) to permit conformational changes of its structure. Gene expression analysis revealed that each gene was associated with distinct temporal expression during turion formation. APL2 and APL3 were highly expressed in earlier stages of turion development, while APL1 expression was reduced throughout turion development. CONCLUSIONS: These results suggest that the differential expression of APLs could be used to enhance energy flow from photosynthesis to storage of carbon in aquatic plants, making duckweeds a useful alternative biofuel feedstock.
Calvino, M., Bruggmann R, Messing J.  2011.  Characterization of the small RNA component of the transcriptome from grain and sweet sorghum stems. BMC Genomics. 12:356. AbstractWebsite
ABSTRACT: BACKGROUND: Sorghum belongs to the tribe of the Andropogoneae that includes potential biofuel crops like switchgrass, Miscanthus and successful biofuel crops like corn and sugarcane. However, from a genomics point of view sorghum has compared to these other species a simpler genome because it lacks the additional rounds of whole genome duplication events. Therefore, it has become possible to generate a high-quality genome sequence. Furthermore, cultivars exists that rival sugarcane in levels of stem sugar so that a genetic approach can be used to investigate which genes are differentially expressed to achieve high levels of stem sugar. RESULTS: Here, we characterized the small RNA component of the transcriptome from grain and sweet sorghum stems, and from F2 plants derived from their cross that segregated for sugar content and flowering time. We found that variation in miR172 and miR395 expression correlated with flowering time whereas variation in miR169 expression correlated with sugar content in stems. Interestingly, genotypic differences in the ratio of miR395 to miR395* were identified, with miR395* species expressed as abundantly as miR395 in sweet sorghum but not in grain sorghum. Finally, we provided experimental evidence for previously annotated miRNAs detecting the expression of 25 miRNA families from the 27 known and discovered 9 new miRNAs candidates in the sorghum genome. CONCLUSIONS: Sequencing the small RNA component of sorghum stem tissue provides us with experimental evidence for previously predicted microRNAs in the sorghum genome and microRNAs with a potential role in stem sugar accumulation and flowering time.
Wang, W, Wu Y, Yan Y, Ermakova M, Kerstetter R, Messing J.  2010.  DNA barcoding of the Lemnaceae, a family of aquatic monocots. BMC Plant Biol. 10:205. AbstractWebsite
BACKGROUND: Members of the aquatic monocot family Lemnaceae (commonly called duckweeds) represent the smallest and fastest growing flowering plants. Their highly reduced morphology and infrequent flowering result in a dearth of characters for distinguishing between the nearly 38 species that exhibit these tiny, closely-related and often morphologically similar features within the same family of plants. RESULTS: We developed a simple and rapid DNA-based molecular identification system for the Lemnaceae based on sequence polymorphisms. We compared the barcoding potential of the seven plastid-markers proposed by the CBOL (Consortium for the Barcode of Life) plant-working group to discriminate species within the land plants in 97 accessions representing 31 species from the family of Lemnaceae. A Lemnaceae-specific set of PCR and sequencing primers were designed for four plastid coding genes (rpoB, rpoC1, rbcL and matK) and three noncoding spacers (atpF-atpH, psbK-psbI and trnH-psbA) based on the Lemna minor chloroplast genome sequence. We assessed the ease of amplification and sequencing for these markers, examined the extent of the barcoding gap between intra- and inter-specific variation by pairwise distances, evaluated successful identifications based on direct sequence comparison of the "best close match" and the construction of a phylogenetic tree. CONCLUSIONS: Based on its reliable amplification, straightforward sequence alignment, and rates of DNA variation between species and within species, we propose that the atpF-atpH noncoding spacer could serve as a universal DNA barcoding marker for species-level identification of duckweeds.
Calviño, M, Miclaus M, Bruggmann R, Messing J.  2009.  Molecular Markers for Sweet Sorghum Based on Microarray Expression Data. Rice. 2:129-142. AbstractWebsite
Using an Affymetrix sugarcane genechip, we previously identified 154 genes differentially expressed between grain and sweet sorghum. Although many of these genes have functions related to sugar and cell wall metabolism, dissection of the trait requires genetic analysis. Therefore, it would be advantageous to use microarray data for generation of genetic markers, shown in other species as single-feature polymorphisms (SFPs). As a test case, we used the GeSNP software to screen for SFPs between grain and sweet sorghum. Based on this screen, out of 58 candidate genes, 30 had single-nucleotide polymorphisms (SNPs) from which 19 had validated SFPs. The degree of nucleotide polymorphism found between grain and sweet sorghum was in the order of one SNP per 248 base pairs, with chromosome 8 being highly polymorphic. Indeed, molecular markers could be developed for a third of the candidate genes, giving us a high rate of return by this method.
Calviño, M, Bruggmann R, Messing J.  2008.  Screen of Genes Linked to High-Sugar Content in Stems by Comparative Genomics. Rice. 1:166-176. AbstractWebsite
One of the great advantages of the fully sequenced rice genome is to serve as a reference for other cereal genomes in particular for identifying genes linked to unique traits. A trait of great interest is reduced lignocellulose in the stem of related species in favor of fermentable sugars as a source of biofuels. While sugarcane is one of the most efficient biofuel crops, little is known about the underlying gene repertoire involved in it. Here, we take advantage of the natural variation of sweet and grain sorghum to uncover genes that are conserved in rice, sorghum, and sugarcane but differently expressed in sweet versus grain sorghum by using a microarray platform and the syntenous alignment of rice and sorghum genomic regions containing these genes. Indeed, enzymes involved in carbohydrate accumulation and those that reduce lignocellulose can be identified.