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烟草液泡pH相关基因NtPH1的CRISPR/Cas9敲除载体的构建
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作者 李嘉敏 汪敏 +4 位作者 马坤 洪晓喻 崔超军 陈君宇 祝钦泷 《植物学研究》 2016年第4期132-138,共7页
植物液泡pH的碱性程度是蓝色花形成的一个重要因素。烟草NtPH1是矮牵牛蓝色花形成相关的控制液泡pH的PhPH1基因的直系同源基因。为了研究其功能,本研究利用植物CRISPR/Cas9系统构建NtPH1的双靶点敲除载体。通过对NtPH1基因片段的序列分... 植物液泡pH的碱性程度是蓝色花形成的一个重要因素。烟草NtPH1是矮牵牛蓝色花形成相关的控制液泡pH的PhPH1基因的直系同源基因。为了研究其功能,本研究利用植物CRISPR/Cas9系统构建NtPH1的双靶点敲除载体。通过对NtPH1基因片段的序列分析,选择两个PAM序列相距60 bp的位点作为靶序列,通过重叠PCR直接把两个靶位点分别拼接到AtU6-29和AtU6-26两个启动子驱动的sgRNA表达盒上,再利用Golden Gate克隆的方法把其连入pYLCRISPR/Cas9Pubi-H载体,重组克隆经菌落PCR筛选、质粒DNA酶切鉴定与测序,获得了NtPH1基因的双靶点敲除载体pCas9-PH1T1T2。这为进一步开展NtPH1基因的功能研究与蓝色花的基因工程应用奠定了基础。 展开更多
关键词 烟草 液泡pH NtPH1 CRISPR/Cas9系统 载体构建
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From Golden Rice to aSTARice:Bioengineering Astaxanthin Biosynthesis in Rice Endosperm 被引量:52
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作者 Qinlong Zhu Dongchang Zeng +8 位作者 Suize Yu chaojun cui Jiamin Li Heying Li Junyu Chen Runzhao Zhang Xiucai Zhao Letian Chen Yao-Guang Liu 《Molecular Plant》 SCIE CAS CSCD 2018年第12期1440-1448,共9页
Carotenoids are important phytonutrients with antioxidant properties,and are widely used in foods and feedstuffs as Supplements.Astaxanthin,a red-colored ketocarotenoid,has strong antioxidant activity and thus can ben... Carotenoids are important phytonutrients with antioxidant properties,and are widely used in foods and feedstuffs as Supplements.Astaxanthin,a red-colored ketocarotenoid,has strong antioxidant activity and thus can benefit human health.However,astaxanthin is not produced in most higher plants.Here we report the bioengineering of astaxanthin biosynthesis in rice endosperm by introducing four synthetic genes,sZmPSY1,sPaCrtl,sCrBKT,and sHpBHY,which encode the enzymes phytoene synthase,phytoene desaturase,β-carotene ketolase,and β-carotene hydroxylase,respectively.Transgneic overexpression of two (sZmPSY1 and sPaCrtl),three (sZmPSY1,sPaCrtl and sCrBKT),and all these four genes driven by rice endosperm-specific promoters established the Carotenoid/ketocarotenoid/astaxanthin biosynthetic pathways in the endosperm and thus resulted in various types of germplasm,from the yellow-grained β-caro- tene-enriched Golden Rice to orange-red-grained Canthaxanthin Rice and Astaxanthin Rice,respectively. Grains Of Astaxanthin Rice were enriched with astaxanthin in the endosperm and had higher antioxidant activity.These results proved that introduction of a minimal set of four transgenes enables de novo biosynthesis of astaxanthin in therice endosperm.This work provides a Successful example for synthetic biology in plants and biofortification in crops;the biofortified rice products generated by this study could be consumed as health-promoting foods and processed tO produce dietary supplements. 展开更多
关键词 BIOFORTIFICATION MULTIGENE metabolic engineering ASTAXANTHIN BIOSYNTHESIS RICE ENDOSPERM
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