Protocol: An improved high-throughput method for generating tissue samples in 96-well format for plant genotyping (Ice-Cap 2.0).
Protocol: An improved high-throughput method for generating tissue samples in 96-well format for plant genotyping (Ice-Cap 2.0).
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方案：一种改进的高通量方法，用于以96孔格式生成植物基因分型的组织样品（ICE-CAP 2.0）。
DOI：
10.1186/1746-4811-3-8
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发表时间：
2007-06-12
期刊：
影响因子：
5.1
通讯作者：
Krysan PJ
中科院分区：
文献类型：
--
作者：
Clark KA;Krysan PJ
我们之前开发了一种名为“Ice-Cap”的高通量系统，用于在 96 孔格式中培养拟南芥幼苗，并快速收集组织以进行后续 DNA 提取和基因分型。虽然最初描述的 Ice-Cap 方法是高通量基因分型的有效工具，但第一版 Ice-Cap 的一个缺点是最佳幼苗生长高度依赖于特定的环境条件。在这里，我们描述了 Ice-Cap 方法的几项技术改进，使其更加稳健，并提供了用于实施该方法的详细协议。 Ice-Cap 2.0 的关键创新是连续浇水系统的开发。浇水系统的添加使得幼苗生长板在整个生长期都可以在没有盖子的情况下进行培养，这反过来又使得幼苗生长比使用原始方法观察到的更加均匀和健壮。我们还确定，在收获组织之前在上下板之间插入木串可以更容易地在冷冻后分离板。使用 Ice-Cap 2.0 方法生长的幼苗在 Ice-Cap 板中的存活时间是使用原始方法生长的幼苗的两倍。我们开发的连续浇水系统为使用最初描述的 Ice-Cap 系统时可能遇到的幼苗生长不理想的问题提供了有效的解决方案。这种新颖的浇水系统和对冰帽程序的一些额外修改提高了该方法的稳健性和实用性。
We previously developed a high-throughput system called 'Ice-Cap' for growing Arabidopsis seedlings in a 96-well format and rapidly collecting tissue for subsequent DNA extraction and genotyping. While the originally described Ice-Cap method is an effective tool for high-throughput genotyping, one shortcoming of the first version of Ice-Cap is that optimal seedling growth is highly dependent on specific environmental conditions. Here we describe several technical improvements to the Ice-Cap method that make it much more robust and provide a detailed protocol for implementing the method. The key innovation underlying Ice-Cap 2.0 is the development of a continuous watering system. The addition of the watering system allows the seedling growth plates to be incubated without a lid for the duration of the growth period, which in turn allows for much more uniform and robust seedling growth than was observed using the original method. We also determined that inserting wooden skewers between the upper and lower plates prior to tissue harvest made it easier to separate the plates following freezing. Seedlings grown using the Ice-Cap 2.0 method remain viable in the Ice-Cap plates twice as long as seedlings grown using the original method. The continuous watering system that we have developed provides an effective solution to the problem of sub-optimal seedling growth that can be encountered when using the originally described Ice-Cap system. This novel watering system and several additional modifications to the Ice-Cap procedure have improved the robustness and utility of the method.
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影响因子：
5.1
作者：
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通讯作者：
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