The isolation of early nuclear endosperm of Oryza sativa to facilitate gene expression analysis and screening imprinted genes

Plant Methods

Meng-xiang Sun¹  Xiongbo Peng¹  Xiaobo Yu¹  Quan Kuang¹ 

[1] College of Life Science, State Key Laboratory of Hybrid Rice, Wuhan University, Wuhan 430072, China

关键词: Gene imprinting;    Parental-of-origin genes;    Endosperm;    Rice;   

Others  :  1230468
DOI  :  10.1186/s13007-015-0092-4

 received in 2015-08-08, accepted in 2015-10-13,  发布年份 2015

【 摘 要 】

Background

Since the quality and yield of rice production depends on endosperm development, previous studies have focused on the molecular mechanism that regulates this developmental process. Recently, how this process is epigenetically regulated has become an important topic. However, the gene expression analysis and screening imprinted genes during early endosperm development remain challenging since the isolation of early endosperm has not been possible. Here, we report a procedure for the isolation of endosperm at 24 or 48 HAP (hours after pollination) during the free nuclear stage of endosperm development.

Results

This technique allows for rapid and convenient collection of pure free nuclear endosperm. Early endosperm RNA can then be extracted from the isolated endosperm cells using dynabeads. Our results showed that the quality of RNA is satisfactory for gene expression analysis and screening the parental-of-origin specific genes in early endosperm.

Conclusions

Thus, we offer a reliable method to overcome one of the major obstacles in the investigation of the molecular mechanisms of early endosperm development. Our approach can be used for accurate gene expression analysis and screening of imprinted genes, and facilitates the confirmation of endosperm-specific gene expression at the very early stages of endosperm development. This method could also be used in other species to collect early free nuclear endosperm.

【 授权许可】

   
2015 Kuang et al.

【 图 表 】

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【 参考文献 】

• [1]Dai XM, Huang QC, Li GP, Hu XM, Qin GY. Observation on the process of double fertilization in rice using laser scanning confocal microscopy. J Henan Agric Sci. 2007; 6:25-30.
• [2]Dai XM, Huang QC, Qin GY. Observation on the double fertilization in rice by using confocal microscopy. Guihaia. 2008; 28:15-19.
• [3]Gu YJ, Xiong F, Wang Z, Chen G, Li WF. A contrast of the endosperm development between rice and wheat. J Nanjing Norm Univ (Nat Sci Edition). 2001; 24:65-74.
• [4]Yang HY. The development of endosperm. In: Rice reproductive biology. Yang HY, editor. Zhejiang University Press, Hangzhou; 2005: p.126-144.
• [5]Wang Z, Gu YJ, Zheng YK, Wang HH. Structure observation of rice endosperm cell development and its mineral element analysis. Chin J Rice Sci. 2012; 26:693-705.
• [6]Olsen OA. Nuclear endosperm development in cereals and Arabidopsis thaliana. Plant Cell. 2004; 16(suppl 1):S214-S227.
• [7]Luo M, Taylor JM, Spriggs A, Zhang HY, Wu XN, Russell S et al.. A genome-wide survey of imprinted genes in rice seeds reveals imprinting primarily occurs in the endosperm. PLoS Genet. 2011; 7(6):e1002125.
• [8]Luo M, Bilodeau P, Dennis ES, Peacock WJ, Chaudhury A. Expression and parent-of-origin effects for FIS2, MEA, and FIE in the endosperm and embryo of developing Arabidopsis seeds. PNAS. 2000; 97(19):10637-10642.
• [9]Luo M, Platten D, Chaudhury A, Peacock WJ, Dennis ES. Expression, imprinting, and evolution of rice homologs of the polycomb group genes. Mol Plant. 2009; 2(4):711-723.
• [10]Li SS, Zhou B, Peng XB, Kuang Q, Huang XL, Yao JL et al.. OsFIE2 plays an essential role in the regulation of rice vegetative and reproductive development. New Phytol. 2014; 201(1):66-79.
• [11]Nallamilli BRR, Zhang J, Mujahid H, Malone BM, Bridges SM, Peng ZH. Polycomb group gene OsFIE2 regulates rice (Oryza sativa) seed development and grain filling via a mechanism distinct from Arabidopsis. PLoS Genet. 2013; 9(3):e1003322.
• [12]Danilevskaya ON, Hermon P, Hantke S, Muszynski MG, Kollipara K, Ananiev EV. Duplicated fie genes in maize expression pattern and imprinting suggest distinct functions. Plant Cell. 2003; 15(2):425-438.
• [13]Conrad LJ, Khanday I, Johnson C, Guiderdoni E, An G, Vijayraghavan U et al.. The polycomb group gene EMF2B is essential for maintenance of floral meristem determinacy in rice. Plant J. 2014; 80(5):883-894.
• [14]Raissig MT, Baroux C, Grossniklaus U. Regulation and flexibility of genomic imprinting during seed development. Plant Cell. 2011; 23(1):16-26.
• [15]Zhang HY, Dong HT, Yao HG, Xiang YW, Tan XL, Yongqi F et al.. Isolation of total RNA from rice endosperm and expression of important genes related to grain quality in rice (Oryza sativa). Chin J Rice Sci. 2004; 19(2):105-110.
• [16]Zhao P, Zhou XM, Zhang LY, Wang W, Ma LG, Yang LB et al.. A bipartite molecular module controls cell death activation in the basal cell lineage of plant embryos. PLoS Biol. 2013; 11(9):e1001655.
• [17]Zhang BT, Carrie C, Ivanova A, Narsai R, Murcha MW, Duncan O et al.. LETM proteins play a role in the accumulation of mitochondrially encoded proteins in Arabidopsis thaliana and AtLETM2 displays parent of origin effects. J Biol Chem. 2012; 287(50):41757-41773.
• [18]Ye RJ, Zhou F, Lin YJ. Two novel positive cis-regulatory elements involved in green tissue-specific promoter activity in rice (Oryza sativa L ssp.). Plant Cell Rep. 2012; 31(7):1159-1172.
• [19]Cai M, Wei J, Li XH, Xu CG, Wang SP. A rice promoter containing both novel positive and negative cis-elements for regulation of green tissue-specific gene expression in transgenic plants. Plant Biotechnol J. 2007; 5(5):664-674.
• [20]Hiei Y, Ohta S, Komari T, Kumashiro T. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J. 1994; 6(2):271-282.
• [21]Cai M, Yuan T, Duan L, Li X, Wang S. Identification of potential protein regulators bound to the tissue-specific positive and negative cis-acting elements of a green tissue-specific promoter in rice. Plant Biol. 2008; 10(6):771-777.
• [22]Lin ZM, Su J, Chen ZJ, Zhou SF, Wang F. Characterization of PsbR3 Promoter of Rice in Photosynthesis. Chin J biochem Mol Biol. 2014; 30(2):179-186.
• [23]Lee DS, Chen LJ, Li CY, Liu YS, Tan XL, Lu BR et al.. The Bsister MADS gene FST determines ovule patterning and development of the zygotic embryo and endosperm. PLoS One. 2013; 8(3):e58748.
• [24]Zhao J, Xin HP, Qu LH, Ning J, Peng XB, Yan TT et al.. Dynamic changes of transcript profiles after fertilization are associated with de novo transcription and maternal elimination in tobacco zygote, and mark the onset of the maternal-to-zygotic transition. Plant J. 2011; 65(1):131-145.
• [25]Tang XC, Liu Y, He YC, Ma LG, Sun MX. Exine dehiscing induces rape microspore polarity, which results in different daughter cell fate and fixes the apical–basal axis of the embryo. J Exp Bot. 2013; 64(1):215-228.
• [26]Ma LG, Xin HP, Qu LH, Zhao J, Yang LB, Zhao P et al.. Transcription profile analysis reveals that zygotic division results in uneven distribution of specific transcripts in apical/basal cells of tobacco. PLoS One. 2011; 6(1):e15971.
• [27]Xin HP, Peng XB, Ning J, Yan TT, Ma LG, Sun MX. Expressed sequence-tag analysis of tobacco sperm cells reveals a unique transcriptional profile and selective persistence of paternal transcripts after fertilization. Sex Plant Reprod. 2011; 24(1):37-46.