Dataset Information

De novo assembly, gene annotation and marker development using Illumina paired-end transcriptome sequences in celery (Apium graveolens L.).

ABSTRACT:

Background

Celery is an increasing popular vegetable species, but limited transcriptome and genomic data hinder the research to it. In addition, a lack of celery molecular markers limits the process of molecular genetic breeding. High-throughput transcriptome sequencing is an efficient method to generate a large transcriptome sequence dataset for gene discovery, molecular marker development and marker-assisted selection breeding.

Principal findings

Celery transcriptomes from four tissues were sequenced using Illumina paired-end sequencing technology. De novo assembling was performed to generate a collection of 42,280 unigenes (average length of 502.6 bp) that represent the first transcriptome of the species. 78.43% and 48.93% of the unigenes had significant similarity with proteins in the National Center for Biotechnology Information (NCBI) non-redundant protein database (Nr) and Swiss-Prot database respectively, and 10,473 (24.77%) unigenes were assigned to Clusters of Orthologous Groups (COG). 21,126 (49.97%) unigenes harboring Interpro domains were annotated, in which 15,409 (36.45%) were assigned to Gene Ontology(GO) categories. Additionally, 7,478 unigenes were mapped onto 228 pathways using the Kyoto Encyclopedia of Genes and Genomes Pathway database (KEGG). Large numbers of simple sequence repeats (SSRs) were indentified, and then the rate of successful amplication and polymorphism were investigated among 31 celery accessions.

Conclusions

This study demonstrates the feasibility of generating a large scale of sequence information by Illumina paired-end sequencing and efficient assembling. Our results provide a valuable resource for celery research. The developed molecular markers are the foundation of further genetic linkage analysis and gene localization, and they will be essential to accelerate the process of breeding.

SUBMITTER: Fu N

PROVIDER: S-EPMC3585167 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Publications

De novo assembly, gene annotation and marker development using Illumina paired-end transcriptome sequences in celery (Apium graveolens L.).

Fu Nan N Wang Qian Q Shen Huo-Lin HL

PloS one 20130228 2

<h4>Background</h4>Celery is an increasing popular vegetable species, but limited transcriptome and genomic data hinder the research to it. In addition, a lack of celery molecular markers limits the process of molecular genetic breeding. High-throughput transcriptome sequencing is an efficient method to generate a large transcriptome sequence dataset for gene discovery, molecular marker development and marker-assisted selection breeding.<h4>Principal findings</h4>Celery transcriptomes from four ...[more]

PMID: 23469050

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Project description:BackgroundCollenchyma serves as a mechanical support tissue for many herbaceous plants. Previous work based on solid-state NMR and immunomicroscopy suggested collenchyma cell walls (CWs) may have similar polysaccharide compositions to those commonly found in eudicotyledon parenchyma walls, but no detailed chemical analysis was available. In this study, compositions and structures of cell wall polysaccharides of peripheral collenchyma from celery petioles were investigated.ResultsThis is the first detailed investigation of the cell wall composition of collenchyma from any plant. Celery petioles were found to elongate throughout their length during early growth, but as they matured elongation was increasingly confined to the upper region, until elongation ceased. Mature, fully elongated, petioles were divided into three equal segments, upper, middle and lower, and peripheral collenchyma strands isolated from each. Cell walls (CWs) were prepared from the strands, which also yielded a HEPES buffer soluble fraction. The CWs were sequentially extracted with CDTA, Na2CO3, 1 M KOH and 4 M KOH. Monosaccharide compositions of the CWs showed that pectin was the most abundant polysaccharide [with homogalacturonan (HG) more abundant than rhamnogalacturonan I (RG-I) and rhamnogalacturonan II (RG-II)], followed by cellulose, and other polysaccharides, mainly xyloglucans, with smaller amounts of heteroxylans and heteromannans. CWs from different segments had similar compositions, but those from the upper segments had slightly more pectin than those from the lower two segments. Further, the pectin in the CWs of the upper segment had a higher degree of methyl esterification than the other segments. In addition to the anticipated water-soluble pectins, the HEPES-soluble fractions surprisingly contained large amounts of heteroxylans. The CDTA and Na2CO3 fractions were rich in HG and RG-I, the 1 M KOH fraction had abundant heteroxylans, the 4 M KOH fraction was rich in xyloglucan and heteromannans, and cellulose was predominant in the final residue. The structures of the xyloglucans, heteroxylans and heteromannans were deduced from the linkage analysis and were similar to those present in most eudicotyledon parenchyma CWs. Cross polarization with magic angle spinning (CP/MAS) NMR spectroscopy showed no apparent difference in the rigid and semi-rigid polysaccharides in the CWs of the three segments. Single-pulse excitation with magic-angle spinning (SPE/MAS) NMR spectroscopy, which detects highly mobile polysaccharides, showed the presence of arabinan, the detailed structure of which varied among the cell walls from the three segments.ConclusionsCelery collenchyma CWs have similar polysaccharide compositions to most eudicotyledon parenchyma CWs. However, celery collenchyma CWs have much higher XG content than celery parenchyma CWs. The degree of methyl esterification of pectin and the structures of the arabinan side chains of RG-I show some variation in the collenchyma CWs from the different segments. Unexpectedly, the HEPES-soluble fraction contained a large amount of heteroxylans.

Project description:BackgroundCollenchyma cells occur widely in eudicotyledons and provide mechanical support for growing organs. At maturity, the cells are elongated and have thick, non-lignified walls, which in celery contain cellulose and pectic polysaccharides, together with xyloglucans and heteroxylans and heteromannans. A previous study suggested that at least some of the collenchyma cell wall in celery is laid down after expansion has stopped and is thus secondary. In the present study, we re-examined this. We used chemical analysis and immunomicroscopy to determine changes in the polysaccharide compositions of these walls during development. Additionally, solid-state NMR spectroscopy was used to examine changes in polysaccharide mobilities during development.ResultsWe showed the collenchyma walls are deposited only during cell expansion, i.e. they are primary walls. During cell-wall development, analytical and immunomicroscopy studies showed that within the pectic polysaccharides there were no overall changes in the proportions of homogalacturonans, but there was a decrease in their methyl esterification. There was also a decrease in the proportions of the (1 → 5)-α-L-arabinan and (1 → 4)-β-D-galactan side chains of rhamnogalacturonan I. The proportions of cellulose increased, and to a lesser extent those of xyloglucans and heteroxylans. Immunomicroscopy showed the homogalacturonans occurred throughout the walls and were most abundant in the middle lamellae and middle lamella junctions. Although the (1 → 4)-β-D-galactans occurred only in the rest of the walls, some of the (1 → 5)-α-L-arabinans also occurred in the middle lamellae and middle lamella junctions. During development, the location of the xyloglucans changed, being confined to the middle lamellae and middle lamella junctions early on, but later occurred throughout the walls. The location of the heteroxylans also changed, occurring mostly in the outer walls in young cells, but were more widely distributed in mature cells. Solid-state NMR spectroscopy showed that particularly cellulose, but also homogalacturonans, decreased in mobility during development.ConclusionsOur studies showed that celery collenchyma cell walls are primary and that during their development the polysaccharides undergo dynamic changes. Changes in the mobilities of cellulose and homogalacturonans were consistent with the cell walls becoming stiffer as expansion ceases.

Project description:BACKGROUND:Illumina sequencing of a marker gene is popular in metagenomic studies. However, Illumina paired-end (PE) reads sometimes cannot be merged into single reads for subsequent analysis. When mergeable PE reads are limited, one can simply use only first reads for taxonomy annotation, but that wastes information in the second reads. Presumably, including second reads should improve taxonomy annotation. However, a rigorous investigation of how best to do this and how much can be gained has not been reported. RESULTS:We evaluated two methods of joining as opposed to merging PE reads into single reads for taxonomy annotation using simulated data with sequencing errors. Our rigorous evaluation involved several top classifiers (RDP classifier, SINTAX, and two alignment-based methods) and realistic benchmark datasets. For most classifiers, read joining ameliorated the impact of sequencing errors and improved the accuracy of taxonomy predictions. For alignment-based top-hit classifiers, rearranging the reference sequences is recommended to avoid improper alignments of joined reads. For word-counting classifiers, joined reads could be compared to the original reference for classification. We also applied read joining to our own real MiSeq PE data of nasal microbiota of asthmatic children. Before joining, trimming low quality bases was necessary for optimizing taxonomy annotation and sequence clustering. We then showed that read joining increased the amount of effective data for taxonomy annotation. Using these joined trimmed reads, we were able to identify two promising bacterial genera that might be associated with asthma exacerbation. CONCLUSIONS:When mergeable PE reads are limited, joining them into single reads for taxonomy annotation is always recommended. Reference sequences may need to be rearranged accordingly depending on the classifier. Read joining also relaxes the constraint on primer selection, and thus may unleash the full capacity of Illumina PE data for taxonomy annotation. Our work provides guidance for fully utilizing PE data of a marker gene when mergeable reads are limited.

Dataset Information

De novo assembly, gene annotation and marker development using Illumina paired-end transcriptome sequences in celery (Apium graveolens L.).

Background

Principal findings

Conclusions

Publications

De novo assembly, gene annotation and marker development using Illumina paired-end transcriptome sequences in celery (Apium graveolens L.).

Similar Datasets

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