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http://dx.doi.org/10.4490/algae.2016.31.5.25

Keeping house: evaluation of housekeeping genes for real-time PCR in the red alga, Bostrychia moritziana (Florideophyceae)  

Shim, Junbo (Department of Biology, Kongju National University)
Shim, Eunyoung (Department of Biology, Kongju National University)
Kim, Gwang Hoon (Department of Biology, Kongju National University)
Han, Jong Won (National Marine Biodiversity Institute of Korea)
Zuccarello, Giuseppe C. (School of Biological Sciences, Victoria University of Wellington)
Publication Information
ALGAE / v.31, no.2, 2016 , pp. 167-174 More about this Journal
Abstract
Biological response of cells to variable conditions should affect the expression level of certain genes. Quantification of these changes in target genes needs stable internal controls. Real-time quantitative polymerase chain reaction (PCR) has traditionally used reference or ‘housekeeping’ genes, that are considered to maintain equal expression in different conditions, to evaluate changes in target genes between samples and experimental conditions. Recent studies showed that some housekeeping genes may vary considerably in certain biological samples. This has not been evaluated in red algae. In order to identify the optimal internal controls for real-time PCR, we studied the expression of eleven commonly used housekeeping genes; elongation factor 1-alpha, glyceraldehyde-3-phosphate dehydrogenase, β-actin, polyubiquitin, 30S ribosomal gene, 60S ribosomal gene, beta-tubulin, alpha-tubulin, translation initiation factor, ubiquitin-conjugating enzyme, and isocitrate dehydrogenase in different life-history stages of Bostrychia moritziana. Our results suggest that glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and 30S ribosomal gene, have the most stable gene expression levels between the different life history stages (male, female, carposporophyte, and tetrasporophyte), while the other genes are not satisfactory as internal controls. These results suggest that the combinations of GAPDH and 30S would be useful as internal controls to assess expression level changes in genes that may control different physiological processes in this organism or that may change in different life history stages. These results may also be useful in other red algal systems.
Keywords
Bostrychia moritziana; gene expression; geNorm; housekeeping genes; NormFinder; red algae; RT-qPCR; validation;
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1 Andersen, C. L., Jensen, J. L. & Ørntoft, T. F. 2004. 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.   DOI
2 Asamizu, E., Kato, T., Sato, S., Nakamura, Y., Kaneko, T. & Tabata, S. 2003. Structural analysis of a Lotus japonicus genome. IV. Sequence features and mapping of seventy-three TAC clones which cover the 7.5 Mb regions of the genome. DNA Res. 10:115-122.   DOI
3 Brodie, J. & Zuccarello, G. C. 2007. Systematics of the species rich algae: red algal classification, phylogeny and speciation. In Hodkinson, T. R. & Parnell, J. A. N. (Eds.) Reconstructing the Tree of Life: Taxonomy and Systematics of Species Rich Taxa. CRC Press, Boca Raton, FL, pp. 324-334.
4 Chan, P. -L., Rose, R. J., Abdul Murad, A. M., Zainal, Z., Low, E. -T. L., Ooi, L. C. -L. O., Ooi, S. -E., Yahya, S. & Singh, R. 2014. Evaluation of reference genes for quantitative real-time PCR in oil palm elite planting materials propagated by tissue culture. PLoS One 9:e99774.   DOI
5 Hellemans, J., Mortier, G., De Paepe, A., Speleman, F. & Vandesompele, J. 2007. qBase relative quantification framework and software for management and automated analysis of real-time quantitative PCR data. Genome Biol. 8:R19.   DOI
6 Hibbeler, S., Scharsack, J. P. & Becker, S. 2008. Housekeeping genes for quantitative expression studies in the three-spined stickleback Gasterosteus aculeatus. BMC Mol. Biol. 9:18.   DOI
7 Kamiya, M. & Kawai, H. 2002. Dependence of the carposporophyte on the maternal gametophyte in three ceramiacean algae (Rhodophyta), with respect to carposporophyte development, spore production and germination success. Phycologia 41:107-115.   DOI
8 Kamiya, M., West, J. A. & Hara, Y. 2011. Induction of apomixis by outcrossing between genetically divergent entities of Caloglossa leprieurii (Ceramiales, Rhodophyta) and evidence of hybrid apomicts in nature. J. Phycol. 47:753-762.   DOI
9 Kim, G. H., Han, J. H., Kim, B., Han, J. W., Nam, S. W., Shin, W., Park, J. W. & Yih, W. 2016. Cryptophyte gene regulation in the kleptoplastidic, karyokleptic ciliate Mesodinium rubrum. Harmful Algae 52:23-33.   DOI
10 Kim, G. H., Shim, J. B., Klochkova, T. A., West, J. A. & Zuccarello, G. C. 2008. The utility of proteomics in algae taxonomy: Bostrychia radicans/B. moritziana (Rhodomelaceae Rhodophyta) as a model study. J. Phycol. 44:1519-1528.   DOI
11 Klochkova, T. A., Kang, S. -H., Cho, G. Y., Pueschel, C. M., West, J. A. & Kim, G. H. 2006. Biology of a terrestrial green alga, Chlorococcum sp. (Chlorococcales, Chlorophyta), collected from the Miruksazi stupa in Korea. Phycologia 45:349-358.   DOI
12 Kowalczyk, N., Rousvoal, S., Hervé, C., Boyen, C. & Collén, J. 2014. RT-qPCR normalization genes in the red alga Chondrus crispus. PLoS One 9:e86574.   DOI
13 Vandesompele, J., De Preter, K., Pattyn, F., Poppe, B., Van Roy, N., De Paepe, A. & Speleman, F. 2002. Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol. 3:Research0034.
14 Mortazavi, A., Williams, B. A., McCue, K., Schaeffer, L. & Wold, B. 2008. Mapping and quantifying mammalian transcriptomes by RNA-Seq. Nat. Methods 5:621-628.   DOI
15 Ransbotyn, V. & Reusch, T. B. H. 2006. Housekeeping gene selection for quantitative real-time PCR assays in the seagrass Zostera marina subjected to heat stress. Limnol. Oceanogr. Methods 4:367-373.   DOI
16 Saunders, G. W. & Hommersand, M. H. 2004. Assessing red algal supraordinal diversity and taxonomy in the context of contemporary systematic data. Am. J. Bot. 91:1494-1507.   DOI
17 West, J. A., Zuccarello, G. C. & Kamiya, M. 2001. Reproductive patterns of Caloglossa species (Delesseriaceae, Rhodophyta) from Australia and New Zealand: multiple origins of asexuality in C. leprieurii. Literature review on apomixis, mixed-phase, bisexuality and sexual compatibility. Phycol. Res. 49:183-200.   DOI
18 Wu, X., Huang, A., Xu, M., Wang, C., Jia, Z., Wang, G. & Niu, J. 2013. Variation of expression levels of seven house-keeping genes at different life-history stages in Porphyra yezoensis. PLoS One 8:e60740.   DOI
19 Yoon, H. S., Hackett, J. D., Ciniglia, C., Pinto, G. & Bhattacharya, D. 2004. A molecular timeline for the origin of photosynthetic eukaryotes. Mol. Biol. Evol. 21:809-818.   DOI
20 Zuccarello, G. C. & West, J. A. 2011. Insights into evolution and speciation in the red alga Bostrychia: 15 years of research. Algae 26:21-32.   DOI