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학술연구/단체지원/교육 등 연구자 활동을 지속하도록 DBpia가 지원하고 있어요.
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논문 기본 정보
- 자료유형
- 학술저널
- 저자정보
- 발행연도
- 2016.10
- 수록면
- 768 - 775 (8page)
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The Arabidopsis female gametophyte contains seven cells with eight haploid nuclei buried within layers of sporophytic tissue. Following double fertilization, the egg and central cells of the gametophyte develop into the embryo and endosperm of the seed, respectively. The epigenetic status of the central cell has long presented an enigma due both to its inaccessibility, and the fascinating epigenome of the endosperm, thought to have been inherited from the central cell following activity of the DEMETER demethylase enzyme, prior to fertilization. Here, we present for the first time, a method to isolate pure populations of Arabidopsis central cell nuclei. Utilizing a protocol designed to isolate leaf mesophyll protoplasts, we systematically optimized each step in order to efficiently separate central cells from the female gametophyte. We use initial manual pistil dis-section followed by the derivation of central cell proto-plasts, during which process the central cell emerges from the micropylar pole of the embryo sac. Then, we use a modified version of the Isolation of Nuclei TAgged in specific Cell Types (INTACT) protocol to purify central cell nuclei, resulting in a purity of 75-90% and a yield sufficient to undertake downstream molecular analyses. We find that the process is highly dependent on the health of the original plant tissue used, and the efficiency of protoplasting solution infiltration into the gametophyte. By isolating pure central cell populations, we have enabled elucidation of the physiology of this rare cell type, which in the future will provide novel insights into Arabidopsis reproduction.
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참고문헌 (43)
참고문헌 신청
Bechtold, N. / 1998 / In planta Agrobacteriummediated transformation of adult Arabidopsis thaliana plants by vacuum infiltration / Methods Mol. Biol 82:259~266
Birnbaum, K. / 2005 / Cell type-specific expression profiling in plants via cell sorting of protoplasts from fluorescent reporter lines / Nat. Methods 2:615~619
Boyes, D. C. / 2001 / Growth stage-based phenotypic analysis of Arabidopsis : a model for high throughput functional genomics in plants / Plant Cell 13:1499~1510
Chen, J. / 2015 / A highly efficient maize nucellus protoplast system for transient gene expression and studying programmed cell death-related processes / Plant Cell Rep 34:1239~1251
Choi, Y. / 2002 / DEMETER, a DNA glycosylase domain protein, is required for endosperm gene imprinting and seed viability in Arabidopsis / Cell 110:33~42
Birnbaum, K. / 2005 / Cell type-specific expression profiling in plants via cell sorting of protoplasts from fluorescent reporter lines / Nat. Methods 2:615~619
Boyes, D. C. / 2001 / Growth stage-based phenotypic analysis of Arabidopsis : a model for high throughput functional genomics in plants / Plant Cell 13:1499~1510
Chen, J. / 2015 / A highly efficient maize nucellus protoplast system for transient gene expression and studying programmed cell death-related processes / Plant Cell Rep 34:1239~1251
Choi, Y. / 2002 / DEMETER, a DNA glycosylase domain protein, is required for endosperm gene imprinting and seed viability in Arabidopsis / Cell 110:33~42
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