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NgAgo 研究者发表注意事项

已有 5616 次阅读 2016-8-4 03:09 |系统分类:观点评述

前两天,我质疑了西班牙的MONTOLIU 教授,同时也发给了 NatureBotech 还有两个研究小组。我说只要有一个重复成功,就是被确认了,不必急于结论。其中 PLOS Synbio 回复表示同意。
昨天看到, PLOS Synbio 的主编 Steven 在网上提了几条建议。我附录在后面。

PLOS Synbio 提到的一个网络调查,其实就是网传的140个实验室未能重复的来源。这个调查任何人都可以匿名去填,现在已经涨到 172 了。所以,网传说什么140个实验室没能重复,这个数字会继续攀升。

目前 NgAgo 事件也就是中国媒体(包括英文版)报道了 -- 毕竟相关研究才开始两个月,结论为时过早。澳洲的BURGIO教授也被推到了前台。他仍在继续实验 NgAgo,鉴于中国媒体的报道,他说他宁愿不要成为中国的名人,也许这会影响一些他的某些研究合作项目 (“I would have prefer not to be famous in China. It might impede some research collaborations and I am not very happy with that.”


Hi, I am a community editor for the PLOS Synbio blog. I've become interested in this for two reasons (1) some of our readers were interested in using NgAgo and (2) i would like to try and work out what is going on and to see if anyone has any pro-tips.
I have been directed here by researchers on twitter in a quest to gather material for a blog piece to raise awareness for those considering the technology. From reading the thread and looking at this survey https://dnascissors.wordpress.com/2016/07/15/results-testing-ngago-genome-editing-system
things are not looking encouraging.
I have also been in contact with Addgene who say they will be posting a blog on their experiences later this week which might be of interest, in their reply the following points were made (probably not new to you but I'll share anyway):
  1. NgAgo uses 5' phosphorylated DNA guides as opposed to the RNA guides used by similar CRISPR-based genome editing technologies. These DNA guides must be directly delivered to the cells of interest and cannot be made from a plasmid like RNA guides can. Any inefficiencies in delivery of these DNA guides will likely lead to lower editing efficiency;

  1. NgAgo is "one-guide-faithful" and can only load a guide DNA as the NgAgo protein is being produced. This means that, if NgAgo protein is translated but there is no guide DNA around for it to load, it will remain unloaded and will not be directed to cleave a target. To mitigate this issue and generate more successfully loaded NgAgo proteins, Gao et. al., 2016  suggest transfecting target cells with guide DNAs multiple times during the course of a single experiment;

  1. Gao et. al., 2016 only demonstrated genome editing in a few mammalian cancer cell lines. Should researchers attempt to use NgAgo in other systems, there may be more optimization required than that documented in the paper;

  1. Because CRISPR has had much longer to develop as a genome editing technology, there are many more resources available to scientists who would like to develop/use gRNAs and delivery systems for their organism of interest. There isn't yet a large amount of data on the best ways to design and deliver guide DNAs for NgAgo.

If anyone would be happy to share their experiences for our blog, or have any additional resources they had point me in the direction of I would love to hear from you. Best


上一篇:NATURE 未启动对韩春雨的调查
下一篇:澳洲 BURGIO 教授未按韩氏流程实验

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