Biologia plantarum 2019, 63:253-261 | DOI: 10.32615/bp.2019.029

Identification of candidate reference genes in tropical bamboos stable across species, tissues, and developmental stages

S. Chakraborty, S. Dutta, P. Biswas, M. Das*
Department of Life Sciences, Presidency University, 700073 Kolkata, India

Bamboo possesses many unique physiological characteristics, but the molecular understanding of many of these processes remains poorly understood till to date. One major reason is unavailability of sufficient sequence and expression data. Selection of suitable reference genes is pivotal to initiate any gene expression analyses. Although, suitable reference genes have been identified in the temperate bamboo Phyllostachys edulis, it has not been done for tropical bamboo. In this study, expression stability of 10 candidate reference genes were investigated in 4 widely grown tropical bamboo species (Bambusa tulda, B. balcooa, B. bambos, and B. vulgaris), different organs (young leaves from flowering and non flowering culms, flag leaf (leaf just below the mature inflorescence), possible flag leaf (leaf covering the immature inflorescence), culm sheath, internode, root, rhizome, and inflorescence bud), different parts (basal, middle, and tip regions of leaf; internodes located in the basal, middle, and tip region of the branch, and developmental stages early, middle, and late inflorescence buds) by using 3 reliable computational tools (geNorm, NormFinder, and RefFinder). A universal single reference gene for normalization of gene expression data was not identified. However, the eukaryotic initiation factor 4α (eIF4α), clathirin adaptor complexes medium subunit (CAC), and nucleotide tract-binding protein (NTB) were found stable in the selected organs across different bamboo species. On the other hand, eIF4α ranked top when different organs and peptidyl prolyl cis-trans isomerase/cyclophilin (CYP), eukaryotic elongation factor 1α (eEF1α) and ubiquitin 5 (UBQ5) ranked top when different developmental stages of B. tulda were analyzed. Taken together, this study not only identifies reference gene/s that are stable across species, organs, and developmental stages of bamboo, but it also assesses the impacts of major contributing factors regulating expression stability of the reference genes.

Keywords: Bambusa balcooa, B. bambos, B. tulda, B. vulgaris, CAC, CYP, eEF1α, eIF4α, NTB, UBQ5

Accepted: December 8, 2018; Prepublished online: December 8, 2018; Published: January 19, 2019Show citation

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Chakraborty, S., Dutta, S., Biswas, P., & Das, M. (2019). Identification of candidate reference genes in tropical bamboos stable across species, tissues, and developmental stages. Biologia plantarum63(1), 253-261. doi: 10.32615/bp.2019.029.
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References

  1. Andersen, C.L., Jensen, J.L., Ørntoft, T.F.: Normalization of real-time quantitative reverse transcription-PCR data: a model-based variance estimation approach to identify genes suited for normalization, applied to bladder and colon cancer data sets. - Cancer Res. 64: 5245-5250, 2004. Go to original source...
  2. Biswas, P., Chakraborty, S., Dutta, S., Pal, A., Das, M.: Bamboo flowering from the perspective of comparative genomics and transcriptomics. - Front. Plant Sci. 7: 1900, 2016.
  3. Bustin, S., Benes, V., Nolan, T., Pfaffl, M.W.: Quantitative real-time RT-PCR - a perspective. - J. mol. Endocrinol. 34: 597-601, 2005. Go to original source...
  4. Chen, X., Mao, Y., Huang, S., Ni, J., Lu, W., Hou, J., Wang, Y., Zhao, W., Li, M., Wang, Q., Wu, L.: Selection of suitable reference genes for quantitative real-time PCR in Sapium sebiferum. - Front Plant Sci. 8:637, 2017. Go to original source...
  5. Clark, L.G., Londoño, X., Ruiz-Sanchez, E: Bamboo taxonomy and habitat. - In: Liese W., Köhl M. (ed.): Bamboo. Tropical Forestry. Vol. 10. Pp. 1-30. Springer, Cham 2015.
  6. Clark, L.G., Judziewicz, E.J., Tyrrel, C.D.L: Aulonemia ximenae (Poaceae: Bambusoideae), a new northern Andean species with fimbriate sheath margin. - Bamboo Sci. Cult. 20: 1-6, 2007.
  7. Clark, L.G., Zhang, W., Wendel, J.F.: A phylogeny of the grass family (Poaceae) based on ndhF sequence data. - Syst. Bot. 20: 436-460, 1995. Go to original source...
  8. Das, M., Bhattacharya, S., Singh, P., Filgueiras, T.S., Pal, A.: Bamboo taxonomy and diversity in the era of molecular markers. - Adv. bot. Res. 47: 225-268, 2008. Go to original source...
  9. Das, M., Haberer, G., Panda, A., Das Laha, S., Ghosh, T.C., Schäffner, A.R.: Expression pattern similarities support the prediction of orthologs retaining common functions after gene duplication events. - Plant Physiol. 171: 2343-2357, 2016. Go to original source...
  10. Das, M., Pal, A.: In vitro regeneration of Bambusa balcooa Roxb.: Factors affecting changes of morphogenetic competence in the axillary buds. - Plant Cell Tissue Organ Cult. 81: 109-112, 2005. Go to original source...
  11. Das, M., Reichman, J.R., Haberer, G., Welzl, G., Aceituno, F.F., Mader, M.T., Watrud, L.S., Pfleeger, T.G., Gutiérrez, R.A., Schäffner, A.R., Olszyk, D.M.: A composite transcriptional signature differentiates responses towards closely related herbicides in Arabidopsis thaliana and Brassica napus. - Plant mol. Biol. 72: 545-56, 2010. Go to original source...
  12. De Andrade, L.M., Dos Santos Brito, M., Fávero Peixoto, R., Jr., Marchiori, P.E.R., Nóbile, P.M., Martins, A.P.B., Ribeiro, R.V., Creste, S.: Reference genes for normalization of qPCR assays in sugarcane plants under water deficit. - Plant Methods 13: 28-37, 2017. Go to original source...
  13. Dutta, S., Biswas, P., Chakraborty, S., Mitra, D., Pal, A., Das, M.: Identification, characterization and gene expression analyses of important flowering genes related to photoperiodic pathway in bamboo. - BMC Genomics 19: 190, 2018. Go to original source...
  14. Fan, C., Ma, J., Guo, Q., Li, X., Wang, H., Lu, M.: Selection of reference genes for quantitative real-time PCR in bamboo (Phyllostachys edulis). - PLoS ONE 8: e56573, 2013. Go to original source...
  15. Fedick, A., Su, J., Jalas, C., Treff, N.R.: High-throughput real-time PCR-based genotyping without DNA purification. - BMC Res Notes 5: 573, 2012. Go to original source...
  16. Gielis, J., Everaert, I., De Loose, M.: Genetic variability and relationships in Phyllostachys using random amplified polymorphic DNA. -In: Chapman, G.P. (ed): TheBbamboos. Pp. 107-124. Academic Press, London 1997.
  17. Ginzinger, D.G.: Gene quantification using real-time quantitative PCR: an emerging technology hits the mainstream. - Exp. Hematol. 30: 503-512, 2002. Go to original source...
  18. Guerriero, G., Legay, S., Hausman, J.F.: Alfalfa cellulose synthase gene expression under abiotic stress: a Hitchhiker's guide to RT-qPCR normalization. - PLoS ONE 9: e103808, 2014. Go to original source...
  19. Guo, J., Ling, H., Wu, Q., Xu, L., Que, Y.: The choice of reference genes for assessing gene expression in sugarcane under salinity and drought stresses. - Sci. Rep. 4: 7042, 2014.
  20. Hu, Y., Chen, H., Luo, C., Dong, L., Zhang, S., He, X., Huang, G.: Selection of reference genes for real-time quantitative PCR studies of kumquat in various tissues and under abiotic stress. - Sci. Hort. 174: 207-216, 2014. Go to original source...
  21. Iskandar, H.M., Simpson, R.S., Casu, R.E., Bonnett, G.D., Maclean, D.J, Manners, J.M.: Comparison of reference genes for quantitative real-time polymerase chain reaction analysis of gene expression in sugarcane. - Plant mol. Biol. Rep. 22: 325-337, 2004. Go to original source...
  22. Jain, M., Nijhawan, A., Tyagi, A.K., Khurana, J.P.: Validation of housekeeping genes as internal control for studying gene expression in rice by quantitative real-time PCR. - Biochem. biophys. Res. Commun. 345: 646-51, 2006. Go to original source...
  23. Janská, A., Hodek, J., Svoboda, P., Zámečník, J., Prášil, I.T., Vlasáková, E., Milella, L., Ovesná, J.: The choice of reference gene set for assessing gene expression in barley (Hordeum vulgare L.) under low temperature and drought stress. - Mol. Genet. Genomics 288: 639-49, 2013. Go to original source...
  24. Janzen, D.H.: Why bamboos wait so long to flower. - Annu. Rev. Ecol. Syst. 7: 347-391, 1976. Go to original source...
  25. Jian, B., Liu, B., Bi, Y., Hou, W., Wu, C., Han, T.: Validation of internal control for gene expression study in soybean by quantitative real-time PCR. - BMC mol. Biol. 9: 59, 2008. Go to original source...
  26. Kellogg, E.A.: Flowering plants, monocots, Poaceae. In: Kubitzki K. (ed.): The Families and Genera of Vascular Plants Vol. XIII. Pp. 1-496. Springer, Cham 2015.
  27. Kim, H.Y., Saha, P., Farcuh, M., Li, B., Sadka, A., Blumwald, E.: RNA-seq analysis of spatio-temporal gene expression patterns during fruit development revealed reference genes for transcript normalization in plums. - Plant mol. Biol. Rep. 33: 1634-1649, 2015. Go to original source...
  28. Li, M.Y., Wang, F., Jiang, Q., Wang, G.L., Tian, C., Xiong, A.S.: Validation and comparison of reference genes for qPCR normalization of celery (Apium graveolens) at different development stages. - Front. Plant Sci. 7: 313, 2016. Go to original source...
  29. Lin, F., Manisseri, C., Fagerström, A., Peck, ML., Vega-Sánchez, ME., William,s B., Chiniquy, DM., Saha, P., Pattathil, S., Conlin, B., Zhu, L., Hahn, MG., Willats, WG., Scheller, HV., Ronald, PC., Bartley, LE.: Cell wall composition and candidate biosynthesis gene expression during rice development. - Plant Cell Physiol. 57: 2058-2075, 2016. Go to original source...
  30. Ling, H., Wu, Q., Guo, J., Xu, L., Que, Y.: Comprehensive selection of reference genes for gene expression normalization in sugarcane by real time quantitative RT-PCR. - Plos ONE 9: e97469, 2014. Go to original source...
  31. Livak, K.J., Schmittgen, T.D.: Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method. - Methods 25: 402-08, 2001. Go to original source...
  32. Lobovikov, M., Paudel, S., Piazza, M., Wu, H.R.: World Bamboo Resources: a Thematic Study Prepared in the Framework of the Global Forest Resources Assessment. - Food and Agriculture Organization of the United Nations, Rome 2007.
  33. Marchesini, V.A., Sala, O.E., Austin, A.T.: Ecological consequences of a massive flowering event of bamboo (Chusqueaculeou) in a temperate forest of Patagonia, Argentina. - J. veg. Sci. 20: 424-432, 2009. Go to original source...
  34. Morgante, C.V., Guimarães, P.M., Martins, A.C.Q., Araújo, A.C.G., Leal-Bertioli, S.C.M., Bertioli, D.J., Brasileiro, A.C.M.: Reference genes for quantitative reverse transcription-polymerase chain reaction expression studies in wild and cultivated peanut. - BMC Res. Notes 4: 339, 2011. Go to original source...
  35. Niu, X., Chen, M., Huang, X., Chen, H., Tao, A., Xu, J., Qi, J.: Reference gene selection for qRT-PCR normalization analysis in kenaf (Hibiscus cannabinus L.) under abiotic stress and hormonal stimuli. - Front. Plant Sci. 8: 771, 2017. Go to original source...
  36. Owen, A.: Bamboo!! Improving island economy and resilience with Guam college students. - J. Marine Island Cult. 4: 65-75, 2015. Go to original source...
  37. Pabuayon, I.M., Yamamoto, N., Trinidad, J.L., Longkumer, T., Raorane, M.L., Kohli, A.: Reference genes for accurate gene expression analyses across different tissues, developmental stages and genotypes in rice for drought tolerance. - Rice 9: 32, 2016. Go to original source...
  38. Peng, Z., Lu, Y., Li, L., Zhao, Q., Feng, Q., Gao, Z., Lu, H., Hu, T., Yao, N., Liu, K., Li, Y., Fan, D., Guo, Y., Li, W., Lu, Y., Weng, Q., Zhou, C., Zhang, L., Huang, T., Zhao, Y., Zhu, C., Liu, X., Yang, X., Wang, T., Miao, K., Zhuang, C., Cao, X., Tang, W., Liu, G., Liu, Y., Chen, J., Liu, Z., Yuan, L., Liu, Z., Huang, X., Lu, T., Fei, B., Ning, Z., Han, B., Jiang, Z.: The draft genome of the fast-growing non-timber forest species moso bamboo (Phyllostachys heterocycla). - Nat. Genet. 45: 456-461, 2013. Go to original source...
  39. Ramanayake, S.M.S.D., Meemaduma, V.N., Weerawardene, T.E.: In vitro shoot proliferation and enhancement of rooting for the large-scale propagation of yellow bamboo (Bambusa vulgaris 'Striata') - Sci. Hort. 110: 109-113, 2006. Go to original source...
  40. Reddy, D.S., Bhatnagar-Mathur, P., Reddy, P.S., Sri Cindhuri, K., Sivaji Ganesh A, Sharma, K.K.: Identification and validation of reference genes and their impact on normalized gene expression studies across cultivated and wild Cicer species. - PLoS ONE 11: e0148451, 2016. Go to original source...
  41. Saha, P., Blumwald, E.: Assessing reference genes for accurate transcript normalization using quantitative real-time PCR in pearl millet [Pennisetum glaucum (L.) R. Br.]. - PLoS ONE 9: e106308, 2014. Go to original source...
  42. Santos, FICD., Marini, N., Santos, RSD., Hoffman, BSF., Alves-Ferreira, M., De Oliveira, AC.: Selection and testing of reference genes for accurate RT-qPCR in rice seedlings under iron toxicity. - PLoS ONE 13: e0193418, 2018. Go to original source...
  43. Sertse, D., Disasa, T., Bekele, K., Alebachew, M., Kebede, Y., Eshete, N., Eshetu, S.: Mass flowering and death of bamboo: a potential threat to biodiversity and livelihoods in Ethiopia. - J. biol. environ. Sci. 1: 16-25, 2011.
  44. Silva, R.L.O., Silva, M.D., Ferreira, Neto, J.R.C., Nardi, C.H., Chabregas, S.M., Burnsquist, W.L., Kahl, S., Benko-Iseppon, A.M., Kido, E.A.: Validation of novel reference genes for reverse transcription quantitative real-time PCR in drought stressed sugarcane. - Sci. World J. 2014: 1-12, 2014. Go to original source...
  45. Silver, N., Best, S., Jiang, J., Thein, S.L.: Selection of housekeeping genes for gene expression studies in human reticulocytes using real-time PCR. - BMC mol. Biol. 7: 33, 2006. Go to original source...
  46. Sood, Y.V., Pande, P.C., Tyagi, S., Payra, I., Kulkarni, N., Kulkarni, A.G.: Quality improvement of paper from bamboo and hardwood furnish through fiber fractionation. - J. sci. indian Res. 64: 299-305, 2005.
  47. Subburaj, S., Zhu, D., Li, X., Hu, Y., Yan, Y.: Molecular characterization and expression profiling of Brachypodium distachyon L. cystatin genes reveal high evolutionary conservation and functional divergence in response to abiotic stress. - Front. Plant Sci. 8: 743, 2017. Go to original source...
  48. Vandesompele, J., Preter, K.D., Pattyn, F., Poppe, B., Roy, N.V., Paepe, A.D., Speleman, F.: Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. - Genome Biol 3: 0034.1-0034.11, 2002. Go to original source...
  49. Wai, N.N., Nanko, H., Murakami, K.: A morphological study on the behavior of bamboo pulp fibers in the beating process. - Wood Sci. Technol. 19: 211-222, 1985. Go to original source...
  50. Wan, H., Yuan, W., Ruan, M., Ye, Q., Wang, R., Li, Z., Zhou, G., Yao, Z., Zhao, J., Liu, S., Yang, Y.: Identification of reference genes for reverse transcription quantitative real-time PCR normalization in pepper (Capsicum annuum L.). - Biochem. biophys. Res. Commun. 416: 24-30, 2011. Go to original source...
  51. Wang, M.L., Li, Q.H., Xin, H.H., Chen, X., Zhu, X.J., Li, X.H.: Reliable reference genes for normalization of gene expression data in tea plants (Camellia sinensis) exposed to metal stresses. - PLoS ONE 12: e0175863, 2017. Go to original source...
  52. Wu, F., Shi, X., Lin, X., Liu, Y., Chong, K., Theissen, G., Meng, Z.: The ABCs of flower development: mutational analysis of AP1/FUL-like genes in rice provides evidence for a homeotic (A)-function in grasses. - Plant J. 89: 310-324, 2017. Go to original source...
  53. Yuan, J.L., Yue, J.J., Gu, X.P., Lin, C.S.: Flowering of woody bamboo in tissue culture systems. - Front. Plant Sci. 8: 1589, 2017. Go to original source...
  54. Zhang, W.: Phylogeny of the grass family (Poaceae) from rpl 16 intron sequence data. - Mol. Phylogenetics Evol. 15: 135-146, 2000. Go to original source...