De novo transcriptome analysis using 454 pyrosequencing of the Himalayan Mayapple, Podophyllum hexandrum

BMC Genomics

Research Article

Sharmila Chattopadhyay¹  Ragini Sinha¹  Riddhi Datta¹  Saptarshi Hazra¹  Dipto Bhattacharyya² 

[1] Plant Biology Laboratory, Drug Development/Diagnostics & Biotechnology Division, CSIR-Indian Institute Chemical Biology, 4 Raja S. C. Mullick Road, 700032, Kolkata, India;Plant Biology Laboratory, Drug Development/Diagnostics & Biotechnology Division, CSIR-Indian Institute Chemical Biology, 4 Raja S. C. Mullick Road, 700032, Kolkata, India;Division of Biotechnology, Chonbuk National University, 79 Gobong-ro, 570-752, Iksan-si, Jeollabuk-do, Republic of Korea;

关键词: 454 pyrosequencing;    Podophyllum hexandrum;    Podophyllotoxin biosynthesis;    Transcriptome;   

DOI  :  10.1186/1471-2164-14-748

 received in 2013-01-04, accepted in 2013-10-19,  发布年份 2013

来源: Springer

【 摘 要 】

BackgroundThe Himalayan or Indian Mayapple (Podophyllum hexandrum Royle) produces podophyllotoxin, which is used in the production of semisynthetic anticancer drugs. High throughput transcriptome sequences or genomic sequence data from the Indian Mayapple are essential for further understanding of the podophyllotoxin biosynthetic pathway.Results454 pyrosequencing of a P. hexandrum cell culture normalized cDNA library generated 2,667,207 raw reads and 1,503,232 high quality reads, with an average read length of 138 bp. The denovo assembly was performed by Newbler using default and optimized parameters. The optimized parameter generated 40, 380 assembled sequences, comprising 12,940 contigs and 27,440 singlets which resulted in better assembly as compared to default parameters. BLASTX analysis resulted in the annotation of 40,380 contigs/singlet using a cut-off value of ≤1E-03. High similarity to Medicago truncatula using optimized parameters and to Populus trichocarpa using default parameters was noted. The Kyoto encyclopedia of genes and genomes (KEGG) analysis using KEGG Automatic Annotation Server (KAAS) combined with domain analysis of the assembled transcripts revealed putative members of secondary metabolism pathways that may be involved in podophyllotoxin biosynthesis. A proposed schematic pathway for phenylpropanoids and podophyllotoxin biosynthesis was generated. Expression profiling was carried out based on fragments per kilobase of exon per million fragments (FPKM). 1036 simple sequence repeats were predicted in the P. hexandrum sequences. Sixty-nine transcripts were mapped to 99 mature and precursor microRNAs from the plant microRNA database. Around 961 transcripts containing transcription factor domains were noted. High performance liquid chromatography analysis showed the peak accumulation of podophyllotoxin in 12-day cell suspension cultures. A comparative qRT-PCR analysis of phenylpropanoid pathway genes identified in the present data was performed to analyze their expression patterns in 12-day cell culture, callus and rhizome.ConclusionsThe present data will help the identification of the potential genes and transcription factors involved in podophyllotoxin biosynthesis in P. hexandrum. The assembled transcripts could serve as potential candidates for marker discovery and conservation, which should form the foundations for future endeavors.

【 授权许可】

Unknown   
© Bhattacharyya et al.; licensee BioMed Central Ltd. 2013. This article is published under license to BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.