Transcriptome profiling of Elettaria cardamomum (L.) Maton (small cardamom).

Elettaria cardamomum (L.) Maton, known as 'queen of spices, is a perennial herbaceous monocot of the family Zingiberaceae, native to southern India. Cardamom is an economically valuable spice crop and used widely in culinary and medicinal purposes. In the present study, using Ion Proton RNA sequencing technology, we performed transcriptome sequencing and de novo transcriptome assembly of a wild and five cultivar genotypes of cardamom. RNA-seq generated a total of 22,811,983 (92 base) and 24,889,197 (75 base) raw reads accounting for approximately 8.21GB and 7.65GB of sequence data for wild and cultivar genotypes of cardamom respectively. The raw data were submitted to SRA database of NCBI under the accession numbers SRX1141272 (wild) and SRX1141276 (cultivars). The raw reads were quality filtered and assembled using MIRA assembler resulted with 112,208 and 264,161contigs having N50 value 616 and 664 for wild and cultivar cardamom respectively. The assembled unigenes were functionally annotated using several databases including PlantCyc for pathway annotation. This work represents the first report on cardamom transcriptome sequencing. In order to generate a comprehensive reference transcriptome, we further assembled the raw reads of wild and cultivar genotypes which might enrich the plant transcriptome database and trigger advanced research in cardamom genomics.

Direct link to deposited data

http://www.ncbi.nlm.nih.gov/sra/SRX1141276[accn]

http://www.ncbi.nlm.nih.gov/sra/SRX1141272[accn]

Introduction

Elettaria cardamomum (L.) Maton or small cardamom, belongs to the family Zingiberaceae, is a potential spice crop due to its unique aromatic flavor and multiple health benefits. Cardamom seeds and fruits are the economically significant parts and effectively used as a traditional medicine, food additive and flavoring agent [1]. The essential oil component extracted from the fruits of cardamom was reported to possess antibacterial, anti-inflammatory, analgesic, and antispasmodic activities [2], [3]. In the cardamom essential oil, the bioactive component d- limonene was reported to possess chemopreventive property towards colon cancer, mammary, lung, liver, skin and stomach cancers in rodents [4], [5], [6]. Hence better understanding of genes and pathways associated with the biosynthesis of the active compounds in cardamom might be beneficial for therapeutic purposes as well as selection of superior genotypes. Since there are no genomic and transcriptome data of cardamom available in any of the publicly available databases, the data we presented here provide information on potential genes coding for enzymes responsible for pharmaceutically valuable bioproducts.

Experimental design, materials and methods

Plant material

Fresh leaf, stem, flower, flower bud and young fruit were collected from wild and five cultivar accessions (including landraces and released varieties) of cardamom growing in the JNTBGRI cardamom conservatory. The tissues were immediately frozen in liquid nitrogen and kept as such until processing.

Total RNA isolation and transcriptome sequencing

Total RNA was isolated from all tissues of cardamom using RNeasy plant mini kit combined with modified CTAB method [7]. Quantification and quality analysis were done using 2100 BioAnalyzer (Agilent Technologies). The tissues of wild cardamom were pooled as one sample and the tissues from all cultivars were pooled as second sample representing wild and cultivar genotypes of cardamom. Two sets of total RNA were purified and cDNA library was constructed using Ion Total RNA seq Kit V2 according to manufacturer's instructions. The purified libraries were submitted to RNA sequencing with the Ion Proton sequencer. RNA seq generated a total of 22,811,983 (92 base) and 24,889,197 (75 base) raw reads accounting for approximately 8.21 and 7.65 GB of sequence data for wild and cultivar genotypes of cardamom respectively.

De novo transcriptome assembly and functional annotation

The raw reads were preprocessed to remove low quality bases, tRNAs and rRNAs. De novo assembly was performed with MIRA (Mimicking Intelligent Read Assembly) program [8] which assembles read pairs from partial path into contigs. The transcriptome assembly generated 112,208 and 264,161 contigs for wild and cultivar genotypes and to generate a comprehensive reference transcriptome a combined assembly was performed which yielded 178,745 contigs (Table 1). We annotated the assembled transcripts to NCBI non redundant (NR) protein database, Swissprot, TrEMBL, pfam, PlantCyc, KOG and TAIR10 databases. We also identified the differential expression analysis of significant genes in wild and cultivar genotypes of cardamom. The information we provided might be useful in molecular marker development, SNP identification, genetic manipulation of specific genes to enhance the production of desired compounds and breeding for developing novel elite cultivars.

Table 1

Read and assembly statistics of cardamom transcriptome.

Plant material	Cultivar	Wild
Total number of raw reads	24,889,197	22,811,983
Total number of bases	1,856,105,873	2,105,322,150
> 20	1,518,466,470	1,719,212,155
GC%	55	55
Read length	73 bp	92 bp
Reads after adapter removal and quality trimming	18,921,421	17,515,399
Total contigs	264,161	112,208
Largest contig	1489	1263
N50 contig size	616	664
Average consensus quality	26	26
Contigs with reads without quality values	0	0
Maximum coverage (total)	24,835	17,774
Total size of assembly	16 MB	43 MB

Conflict of interest

The authors declare that they have no competing interests.

Specifications
Organism/cell line/tissue	Wild and cultivar Elettaria cardamomum (L.) Maton
Sex	Monoecious
Sequencer or array type	Ion Proton System™
Data format	Raw data in BAM file
Experimental factors	Transcriptome sequencing of tissues of wild and cultivar cardamom
Experimental features	Freshly collected leaf, stem, flower, flower buds and young fruits were used for RNA isolation, tissues were pooled, RNA seq libraries representing two genotypes of cardamom were sequenced and de novo assembled
Consent	NA
Sample source location	JNTBGRI, Cardamom germplasm conservatory