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. 2014 Jan 23;9(1):e86039.
doi: 10.1371/journal.pone.0086039. eCollection 2014.

Comprehensive transcriptome assembly of Chickpea (Cicer arietinum L.) using sanger and next generation sequencing platforms: development and applications

Himabindu Kudapa  1 , Sarwar Azam  1 , Andrew G Sharpe  2 , Bunyamin Taran  3 , Rong Li  2 , Benjamin Deonovic  4 , Connor Cameron  5 , Andrew D Farmer  5 , Steven B Cannon  6 , Rajeev K Varshney  7
Affiliations

Comprehensive transcriptome assembly of Chickpea (Cicer arietinum L.) using sanger and next generation sequencing platforms: development and applications

Himabindu Kudapa et al. PLoS One. .

Abstract

A comprehensive transcriptome assembly of chickpea has been developed using 134.95 million Illumina single-end reads, 7.12 million single-end FLX/454 reads and 139,214 Sanger expressed sequence tags (ESTs) from >17 genotypes. This hybrid transcriptome assembly, referred to as Cicer arietinumTranscriptome Assembly version 2 (CaTA v2, available at http://data.comparative-legumes.org/transcriptomes/cicar/lista_cicar-201201), comprising 46,369 transcript assembly contigs (TACs) has an N50 length of 1,726 bp and a maximum contig size of 15,644 bp. Putative functions were determined for 32,869 (70.8%) of the TACs and gene ontology assignments were determined for 21,471 (46.3%). The new transcriptome assembly was compared with the previously available chickpea transcriptome assemblies as well as to the chickpea genome. Comparative analysis of CaTA v2 against transcriptomes of three legumes - Medicago, soybean and common bean, resulted in 27,771 TACs common to all three legumes indicating strong conservation of genes across legumes. CaTA v2 was also used for identification of simple sequence repeats (SSRs) and intron spanning regions (ISRs) for developing molecular markers. ISRs were identified by aligning TACs to the Medicago genome, and their putative mapping positions at chromosomal level were identified using transcript map of chickpea. Primer pairs were designed for 4,990 ISRs, each representing a single contig for which predicted positions are inferred and distributed across eight linkage groups. A subset of randomly selected ISRs representing all eight chickpea linkage groups were validated on five chickpea genotypes and showed 20% polymorphism with average polymorphic information content (PIC) of 0.27. In summary, the hybrid transcriptome assembly developed and novel markers identified can be used for a variety of applications such as gene discovery, marker-trait association, diversity analysis etc., to advance genetics research and breeding applications in chickpea and other related legumes.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1
Figure 1. Functional categorization of chickpea Transcript Assembly Contigs (TACs) of the CaTA v2.
Chickpea TACs representing the distribution of genes based on their annotations to terms in the GO were categorized hierarchically according to three principal gene ontologies, viz. biological processes, molecular functions and cellular components. The number of TACs representing each subcategory is shown in Y-axis.
Figure 2
Figure 2. Enzyme classification of chickpea Transcript Assembly Contigs (TACs) among the six enzyme classes.
The graph displays the proportion of genes belonging to each enzyme class.
Figure 3
Figure 3. Distribution of chickpea transcripts in different transcription factor (TF) families.
Based on conserved domain annotation, Transcript Assembly Contigs (TACs) showing significant annotation to transcription factors were classified.
Figure 4
Figure 4. A sample view of chickpea TACs, markers and candidate ISR markers onto Medicago Genome sequence.
This image is from Legume Information System (LIS) GBrowse viewer at http://medtr.comparative-legumes.org/gb2/gbrowse/3.5.1/, shows 1 Mb (17,648,842.18,648,841 of Medicago, chromosome Mt1). Red: There was at least one additional reported CaTA v2 alignment Green: There were no other reported alignments.
See this image and copyright information in PMC

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Grants and funding

The authors are thankful to the DST-INSPIRE program by Department of Science and Technology, Government of India, Indo-German Science Technology Centre (IGSTC), CGIAR Generation Challenge Programme (GCP) and the Saskatchewan Agriculture Development Fund (ADF) for financial support to undertake part of research presented in this study. This work has been undertaken as part of the CGIAR Research Program on Grain Legumes. ICRISAT is a member of CGIAR Consortium. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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