2012年10月9日 訊 /生物谷BIOON/ --在一項(xiàng)新的研究中,，來自美國Iowa State University的Ying Wang和Jeffrey Essner利用一種被稱作轉(zhuǎn)錄激活子樣效應(yīng)物核酸酶(transcription activator-like effector nucleases, TALENs)的定點(diǎn)分子剪刀在對(duì)斑馬魚進(jìn)行基因編輯中取得突破，從而有助于科學(xué)家們?cè)诨畹挠袡C(jī)體中進(jìn)行位點(diǎn)特異性的突變和基因編輯,。相關(guān)研究結(jié)果于2012年發(fā)表在Nature期刊上,，論文標(biāo)題為“In vivo genome editing using high-efficiency TALEN systems”,。論文通信作者是分子生物學(xué)家Stephen Ekker。
通過這種TALENs系統(tǒng),，研究人員首次能夠切掉斑馬的部分DNA,，并插入人工制造的替代DNA序列。這就允許研究人員去激活和可能激活靶基因,，這可能有助于科學(xué)家們研究早期胚胎發(fā)育和成年生活的遺傳特征,。Ying Wang說，這些創(chuàng)新性發(fā)現(xiàn)可能在農(nóng)業(yè)和人類疾病中有著極大的應(yīng)用,。
Wang說,，“這項(xiàng)研究在很多方面都帶來深刻的影響。它可能導(dǎo)致農(nóng)作物和生產(chǎn)性動(dòng)物更加有效地抵抗疾病,，而且它有潛力讓人們對(duì)人類遺傳特征和人類疾病產(chǎn)生新的認(rèn)識(shí),。”
Jeffrey Essner說，在遺傳組成上,，斑馬魚與人類擁有幾乎90%的相同性,，因此研究它們的發(fā)育能夠有助于認(rèn)識(shí)人類疾病。在早期的胚胎發(fā)育階段,，這尤其如此,。
Essner說，“大多數(shù)癌癥是因?yàn)榛螂S著時(shí)間的推移而發(fā)生的零星突變導(dǎo)致的,。在這項(xiàng)研究中,，我們所做的就是讓人們可能能夠操縱這些基因，甚至在成人體內(nèi)也能如此,。”
激活斑馬魚基因的能力可能導(dǎo)致人們?cè)谥委煱┌Y以及人類與動(dòng)物的一系列其他疾病方面取得突破,。
在這項(xiàng)研究中，研究人員能夠更加有效地利用TALENs技術(shù)和準(zhǔn)確地進(jìn)行基因編輯,。在此之前,，Adam Bogdanove教授與Dan Voytas教授、Bing Yang助理教授和Martin Spalding教授在TALENs技術(shù)上作出了巨大的基礎(chǔ)性研究工作,。
從那時(shí)起,，人們就一直利用TALENs技術(shù)來操縱細(xì)菌的遺傳機(jī)制,，但是Essner說,，他們是首次利用TALENs在體內(nèi)進(jìn)行基因編輯，即在活的有機(jī)體體內(nèi)進(jìn)行操作,。
Essner說,，他們想進(jìn)一步開展研究以便繼續(xù)優(yōu)化和完善這種基因編輯技術(shù)。(生物谷Bioon.com)
doi: 10.1038/nature11537
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PMID:
In vivo genome editing using a high-efficiency TALEN system
Victoria M. Bedell, Ying Wang, Jarryd M. Campbell, Tanya L. Poshusta, Colby G. Starker, Randall G. Krug II, Wenfang Tan, Sumedha G. Penheiter, Alvin C. Ma, Anskar Y. H. Leung, Scott C. Fahrenkrug, Daniel F. Carlson, Daniel F. Voytas, Karl J. Clark, Jeffrey J. Essner & Stephen C. Ekker
The zebrafish (Danio rerio) is increasingly being used to study basic vertebrate biology and human disease with a rich array of in vivo genetic and molecular tools. However, the inability to readily modify the genome in a targeted fashion has been a bottleneck in the field. Here we show that improvements in artificial transcription activator-like effector nucleases (TALENs) provide a powerful new approach for targeted zebrafish genome editing and functional genomic applications. Using the GoldyTALEN modified scaffold and zebrafish delivery system, we show that this enhanced TALEN toolkit has a high efficiency in inducing locus-specific DNA breaks in somatic and germline tissues. At some loci, this efficacy approaches 100%, including biallelic conversion in somatic tissues that mimics phenotypes seen using morpholino-based targeted gene knockdowns6. With this updated TALEN system, we successfully used single-stranded DNA oligonucleotides to precisely modify sequences at predefined locations in the zebrafish genome through homology-directed repair, including the introduction of a custom-designed EcoRV site and a modified loxP (mloxP) sequence into somatic tissue in vivo. We further show successful germline transmission of both EcoRV and mloxP engineered chromosomes. This combined approach offers the potential to model genetic variation as well as to generate targeted conditional alleles.
