Protoplast isolation prior to flow cytometry reveals clear patterns of endoreduplication in potato tubers, related species, and some starchy root crops.
Protoplast isolation prior to flow cytometry reveals clear patterns of endoreduplication in potato tubers, related species, and some starchy root crops.
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DOI：
10.1186/s13007-017-0177-3
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发表时间：
2017
期刊：
影响因子：
5.1
通讯作者：
Veilleux RE
中科院分区：
文献类型：
--
作者：
Laimbeer FPE;Holt SH;Makris M;Hardigan MA;Robin Buell C;Veilleux RE
内复制是在没有细胞分裂的情况下DNA复制的过程，与特殊的细胞功能和细胞大小增加有关。控制番茄果实内复制的基因已被证明影响成熟果实的大小。为了研究它在植物器官发育中的作用，需要一种有效的方法来估计内复制。流式细胞术经常被用来评估内复制，但一些组织和物种，其中包括茄子块茎，仍然难以用于流式细胞术的常规组织制备。我们的目标是通过在流式细胞术之前使用原生质体提取来开发一种方法，专门用于评估马铃薯块茎中的内复制。我们提出了一种评估马铃薯(Solanum Tuberosum)块茎组织内复制的方法。我们对这种方法进行了评估，观察到不同品种间块茎髓和皮层的差异一致，但与整个块茎的大小没有明显的关系。此外，我们能够观察到20个野生马铃薯近缘中的16个具有不同的内重复模式，平均内重复指数(EI)在每细胞0.94到2.62个内圈之间。该方案也被应用于一组淀粉类根茎作物品种，虽然只有五种作物中的两种产生了可靠的流动直方图，但这两种作物(红薯和萝卜)的EIS明显低于野生和栽培马铃薯。本文报告的方案已被证明对各种马铃薯品种和相关物种的块茎以及其他淀粉作物的储藏根有效。这种方法为研究马铃薯的形态和发育提供了一个重要的工具，同时揭示了内复制的自然变异，这可能具有农业意义。本文的在线版本(doi：10.1186/s13007-01777-3)包含补充材料，授权用户可以使用。
Endoreduplication, the process of DNA replication in the absence of cell division, is associated with specialized cellular function and increased cell size. Genes controlling endoreduplication in tomato fruit have been shown to affect mature fruit size. An efficient method of estimating endoreduplication is required to study its role in plant organ development. Flow cytometry is often utilized to evaluate endoreduplication, yet some tissues and species, among them the tubers of Solanum tuberosum, remain intractable to routine tissue preparation for flow cytometry. We aimed to develop a method through the use of protoplast extraction preceding flow cytometry, specifically for the assessment of endoreduplication in potato tubers. We present a method for appraising endoreduplication in potato (Solanum tuberosum) tuber tissues. We evaluated this method and observed consistent differences between pith and cortex of tubers and between different cultivars, but no apparent relationship with whole tuber size. Furthermore, we were able to observe distinct patterns of endoreduplication in 16 of 20 wild potato relatives, with mean endoreduplication index (EI) ranging from 0.94 to 2.62 endocycles per cell. The protocol was also applied to a panel of starchy root crop species and, while only two of five yielded reliable flow histograms, the two (sweet potato and turnip) exhibited substantially lower EIs than wild and cultivated potato accessions. The protocol reported herein has proven effective on tubers of a variety of potato cultivars and related species, as well as storage roots of other starchy crops. This method provides an important tool for the study of potato morphology and development while revealing natural variation for endoreduplication which may have agricultural relevance. The online version of this article (doi:10.1186/s13007-017-0177-3) contains supplementary material, which is available to authorized users.