Functional characterisation of banana (Musa spp.) 2-oxoglutarate-dependent dioxygenases involved in flavonoid biosynthesis

5openInternationalInternational coauthor/editorBananas (Musa) are non-grass, monocotyledonous, perennial plants that are well known for their edible fruits. Their cultivation provides food security and employment opportunities in many countries. Banana fruits contain high levels of minerals and phytochemicals, including flavonoids, which are beneficial for human nutrition. To broaden the knowledge on flavonoid biosynthesis in this major crop plant, we aimed to identify and functionally characterise selected structural genes encoding 2-oxoglutarate-dependent dioxygenases, involved in the formation of the flavonoid aglycon. Musa candidates genes predicted to encode flavanone 3-hydroxylase (F3H), flavonol synthase (FLS) and anthocyanidin synthase (ANS) were assayed. Enzymatic functionalities of the recombinant proteins were confirmed in vivo using bioconversion assays. Moreover, transgenic analyses in corresponding Arabidopsis thaliana mutants showed that MusaF3H, MusaFLS and MusaANS were able to complement the respective loss-of-function phenotypes, thus verifying functionality of the enzymes in planta. Knowledge gained from this work provides a new aspect for further research towards genetic engineering of flavonoid biosynthesis in banana fruits to increase their antioxidant activity and nutritional value.openBusche, M.; Acatay, C.; Martens, S.; Weisshaar, B.; Stracke, R.Busche, M.; Acatay, C.; Martens, S.; Weisshaar, B.; Stracke, R

A <i>De Novo</i> Genome Sequence Assembly of the <i>Arabidopsis thaliana</i> Accession Niederzenz-1 Displays Presence/Absence Variation and Strong Synteny

<div><p><i>Arabidopsis thaliana</i> is the most important model organism for fundamental plant biology. The genome diversity of different accessions of this species has been intensively studied, for example in the 1001 genome project which led to the identification of many small nucleotide polymorphisms (SNPs) and small insertions and deletions (InDels). In addition, presence/absence variation (PAV), copy number variation (CNV) and mobile genetic elements contribute to genomic differences between <i>A</i>. <i>thaliana</i> accessions. To address larger genome rearrangements between the <i>A</i>. <i>thaliana</i> reference accession Columbia-0 (Col-0) and another accession of about average distance to Col-0, we created a <i>de novo</i> next generation sequencing (NGS)-based assembly from the accession Niederzenz-1 (Nd-1). The result was evaluated with respect to assembly strategy and synteny to Col-0. We provide a high quality genome sequence of the <i>A</i>. <i>thaliana</i> accession (Nd-1, LXSY01000000). The assembly displays an N50 of 0.590 Mbp and covers 99% of the Col-0 reference sequence. Scaffolds from the <i>de novo</i> assembly were positioned on the basis of sequence similarity to the reference. Errors in this automatic scaffold anchoring were manually corrected based on analyzing reciprocal best BLAST hits (RBHs) of genes. Comparison of the final Nd-1 assembly to the reference revealed duplications and deletions (PAV). We identified 826 insertions and 746 deletions in Nd-1. Randomly selected candidates of PAV were experimentally validated. Our Nd-1 <i>de novo</i> assembly allowed reliable identification of larger genic and intergenic variants, which was difficult or error-prone by short read mapping approaches alone. While overall sequence similarity as well as synteny is very high, we detected short and larger (affecting more than 100 bp) differences between Col-0 and Nd-1 based on bi-directional comparisons. The <i>de novo</i> assembly provided here and additional assemblies that will certainly be published in the future will allow to describe the pan-genome of <i>A</i>. <i>thaliana</i>.</p></div

Assembly statistics.

<p>Metrics of the Nd-1 genome sequence assembly before and after application of SSPACE, GapFiller and subsequent RBH-based manual improvement.</p

Summary of the sizes of large insertions, deletions and HDRs.

<p>The data were compiled from reciprocal read mapping of Nd-1 reads to the Col-0 genome sequence and vice versa. However, the table presents the results regarding PAV from the view of Nd-1; an insertion in Nd-1 is at the same time a deletion in Col-0, and a deletion in Nd-1 is at the same time an insertion in Col-0.</p

Mapping of Nd-1 scaffolds to Col-0 reference sequence.

<p>Schematic chromosomes are shown in grey with centromere positions in purple. Below each chromosome, red bars indicate the frequency of scaffolds. Above each chromosome, black bars show the abundance of the 180 bp centromeric repeat that has been shown to be a major component of <i>A</i>. <i>thaliana</i> centromeric DNA [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0164321#pone.0164321.ref062" target="_blank">62</a>]. Data were calculated for a window size of 50 kbp.</p

Differential regulation of closely related R2R3-MYB transcription factors controls flavonol accumulation in different parts of the seedling-5

<p><b>Copyright information:</b></p><p>Taken from "Differential regulation of closely related R2R3-MYB transcription factors controls flavonol accumulation in different parts of the seedling"</p><p></p><p>The Plant Journal 2007;50(4):660-677.</p><p>Published online Jan 2007</p><p>PMCID:PMC1976380.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p

Differential regulation of closely related R2R3-MYB transcription factors controls flavonol accumulation in different parts of the seedling-3

<p><b>Copyright information:</b></p><p>Taken from "Differential regulation of closely related R2R3-MYB transcription factors controls flavonol accumulation in different parts of the seedling"</p><p></p><p>The Plant Journal 2007;50(4):660-677.</p><p>Published online Jan 2007</p><p>PMCID:PMC1976380.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p

Differential regulation of closely related R2R3-MYB transcription factors controls flavonol accumulation in different parts of the seedling-2

<p><b>Copyright information:</b></p><p>Taken from "Differential regulation of closely related R2R3-MYB transcription factors controls flavonol accumulation in different parts of the seedling"</p><p></p><p>The Plant Journal 2007;50(4):660-677.</p><p>Published online Jan 2007</p><p>PMCID:PMC1976380.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p

A systematic survey in of transcription factors that modulate circadian parameters-3

�5 days. Representative traces of rhythmic expression of oxplants (pink squares) and wild-type (blue circles) are shown. (, ) -ox, (, ) -ox. (, ) :, (, ) :.<p><b>Copyright information:</b></p><p>Taken from "A systematic survey in of transcription factors that modulate circadian parameters"</p><p>http://www.biomedcentral.com/1471-2164/9/182</p><p>BMC Genomics 2008;9():182-182.</p><p>Published online 21 Apr 2008</p><p>PMCID:PMC2410138.</p><p></p

A systematic survey in of transcription factors that modulate circadian parameters-1

��28). (, , ) Representative traces of rhythmic leaf movement of wild-type (blue circles) and ox-lines (other colored symbols) are shown. (, ) The phase angles normalized to a 24-h cycle (CT phase) are plotted with relative amplitude errors (RAE), which indicate the robustness of the rhythm (the lower the RAE the more robust the rhythm). The center of the circle represents a high RAE (= 1). (, ) -ox, (, ) , and () .<p><b>Copyright information:</b></p><p>Taken from "A systematic survey in of transcription factors that modulate circadian parameters"</p><p>http://www.biomedcentral.com/1471-2164/9/182</p><p>BMC Genomics 2008;9():182-182.</p><p>Published online 21 Apr 2008</p><p>PMCID:PMC2410138.</p><p></p