| Determining Genome Size from Spores of Seedless Vascular Plants
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Author:
Date:
2017-06-05
[Abstract] Seedless vascular plants, including ferns and lycophytes, produce spores to initiate the gametophyte stage and to complete sexual reproduction. Approximately 10% of them are apomictic through the production of genomic unreduced spores. Being able to measure the spore nuclear DNA content is therefore important to infer their reproduction mode. Here we present a protocol of spore flow cytometry that allows an efficient determination of the reproductive modes of seedless vascular plants.
[摘要] 无核维管植物，包括蕨类植物和lycophytes，产生孢子，以启动配子体阶段并完成有性繁殖。其中约10％通过生产基因组未还原的孢子而脱水。因此，能够测量孢子核DNA含量对推断其再生模式是重要的。在这里，我们提出了一个孢子流式细胞术方案，可以有效地确定无核维管植物的繁殖模式。
背景 在无核维管植物中，传统上用于推断核DNA含量以及繁殖模式的孢子发生特征，如减数分裂染色体计数。然而，这些方法是耗时的，或只能提供间接证据。 Kuo等人（2017）尚未建立一种估计这些植物的孢子核DNA含量的有效可靠的方法。在这里，我们描述了使用流式细胞术基于Kuo等人（2017）的工作来评估这些植物的孢子基因组大小的方案。
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| Total RNA Extraction from Grape Berry Skin for Quantitative Reverse Transcription PCR and Microarray Analysis
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Author:
Date:
2016-04-05
[Abstract] Extraction of high quality RNA is an essential step for quantitative reverse transcription PCR (qRT-PCR) and microarray analysis. However, it is not easy to extract high quality RNA from fruit materials, which contain high amounts of polysaccharides, lipids and secondary metabolites. Wan and Wilkins (1994) had developed ‘Hot Borate Method’ to isolate high quality RNA. Here, we describe a modified protocol of the ‘Hot Borate Method’ to isolate high quality RNA from grape berry skin for qRT-PCR and microarray analysis (Suzuki et al., 2015a; Suzuki et al., 2015b).
[摘要] 高质量RNA的提取是定量逆转录PCR（qRT-PCR）和微阵列分析的必要步骤。 然而，从含有大量多糖，脂质和次级代谢物的水果材料中提取高质量RNA是不容易的。 Wan和Wilkins（1994）开发了"热硼酸盐方法"来分离高质量的RNA。 在这里，我们描述了"热硼酸盐方法"的修饰的协议，从葡萄浆果皮肤中分离高质量的RNA用于qRT-PCR和微阵列分析（Suzuki等人，2015a; Suzuki等人 al。，2015b）。
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| Isolation of Tonoplast Vesicles from Tomato Fruit Pericarp
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Author:
Date:
2015-12-20
[Abstract] This protocol describes the isolation of tonoplast vesicles from tomato fruit. The vesicles isolated using this procedure are of sufficiently high purity for downstream proteomic analysis whilst remaining transport competent for functional assays. The methodology was used to study the transport of amino acids during tomato fruit ripening (Snowden et al., 2015) and based on the procedure used by Betty and Smith (Bettey and Smith, 1993). Such vesicles may be useful in further studies into the dynamic transfer of metabolites across the tonoplast for storage and metabolism during tomato fruit development.
[摘要] 这个协议描述了从番茄果实隔离tonoplast囊泡。 使用该程序分离的囊泡具有足够高的纯度用于下游蛋白质组分析，同时保留用于功能测定的转运感受态。 该方法用于研究番茄果实成熟期间氨基酸的转运（Snowden等人，2015）并且基于Betty和Smith使用的程序（Bettey和Smith，1993）。 这样的囊泡可用于进一步研究代谢物在整个质膜上的动态转移，用于番茄果实发育期间的储存和代谢。
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