Abstract
Mesua ferrea L. is a multipurpose versatile tree that is well known as a prospective feedstock biodiesel plant. The potentiality of M. ferrea as a sustainable source of feedstock for biodiesel industry is dependent on an extensive knowledge of the genome structure of the plant. However, to date, there has been no genomic research aimed at the exploitation of the biotechnological potential of this species. Flow cytometry with propidium iodide as the DNA stain was used to estimate the nuclear DNA content of Mesua and the 2C value is estimated to be 1.40 ± 0.02 pg. Somatic chromosome count from root-tip cells is found to be 2n = 30 corresponding to the diploid level (n = 15). Fold variation in genome size (1.14) is observed among the plants collected from different geographic locations in Assam and is attributed to reverse transcriptase-RNase H (RT-RH) domains of the Ty1-copia retrotransposons. Dot blot analysis revealed that Ty1-copia accounts for 2.5% of total haploid nuclear genome of Mesua and phylogenetic analyses showed that the RT-RH sequences are heterogeneous and resolved into 3 distinct groups. These results contribute to our understanding about genome organization of Mesua and will provide valuable information for its utilization in future.
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Acknowledgements
RD and RGS thank Ministry of Human Resources Development (MHRD), Government of India for student fellowship. Thanks also to Department of Biosciences and Bioengineering, IITG for instrumentation facility.
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Conceived and designed the experiments: LR, RD, RGS, SM Performed the experiments: RD, RGS Analyzed the data: RD, RGS, SM Contributed reagents/materials/analysis tools: LR Manuscript preparation: LR, RD, RGS.
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Fig. A1: Dot plot of PI-stained gated nuclei of M. ferrea using P. pinnata as standard; (a) external standardization (b) internal standardization and (c) pseudo-internal standardization. Fig. A2: Correlation between genome size and geographical distribution of M. ferrea in Assam, India. Fig. A3: Somatic metaphase chromosome in different individuals of M. ferrea. Dispur (a); Tezpur (b); Jorhat (c); Dibrugarh (d); Silchar (e); Tinsukia (f); Kokrajhar (g); Halflong (h); IIT Guwahati campus (i). Fig. A4: Multiple sequence alignment of the nucleotide sequences corresponding to the RT-RH domain of the Ty1-copia element in M. ferrea clones using ClustalW. Shading was performed by Multiple Sequence Alignment Editor and Shading Utility (Genedoc, version 2.6.002: www.psc.edu/biomed/genedoc) in conservation mode. Darkest shading indicates residues that are 100 percent conserved and are identical in all sequences; lighter shading denotes residues that are identical in most of sequences or functionally identical in all of them. Clones are numbered as MFTY 1 to MFTY-18. Fig. A5: Dot blot hybridization conducted for the determination of Ty1-copia copy number in M. ferrea. (a) Different concentrations of RT-RH PCR product and (b) M. ferrea genomic DNA were serially diluted on row and dot spotted on a membrane (DOC 3290 kb)
Table A1
Estimation of mean 2C nuclear DNA (pg) in ecotypes of M. ferrea using P. pinnata as reference standard (DOC 38 kb)
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Das, R., Shelke, R.G., Rangan, L. et al. Estimation of nuclear genome size and characterization of Ty1-copia like LTR retrotransposon in Mesua ferrea L.. J. Plant Biochem. Biotechnol. 27, 478–487 (2018). https://doi.org/10.1007/s13562-018-0457-7
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DOI: https://doi.org/10.1007/s13562-018-0457-7