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中国生物工程杂志

China Biotechnology
China Biotechnology  2015, Vol. 35 Issue (9): 14-20    DOI: 10.13523/j.cb.20150903
    
Optimization, Synthesis and Transient Expression of Nattokinase Gene in Tobacco (Nicotiana tabacum L.) Leaves
HAN Lan1,2, JU Lin-fang1, NIU Yi-ding1, WANG Ying-chun1, HASI Agula1
1 College of Life Sciences, Inner Mongolia University, Inner Mongolia Key Laboratory of Herbage & Endemic Crop Biotechnology, Hohhot 010021, China;
2 Department of Biochemical engineering, Hohhot Vocational College, Hohhot 010051, China
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Abstract  

The nattokinase encoding gene (sNK) was synthesized by modifying its sequence based on the optimized codon usage in the plant. The first intron of tomato fruit-specific expression E8 gene was inserted in sNK gene to construct sNKi gene by overla Pextension PCR method. These two synthetic nattokinase gene were transiently expressed in tobacco NC89 leaves by agroinfiltration. Real-time quantitative reverse transcription PCR (RT-qPCR) result showed that higher expression level was detected in tobacco leaves which infiltrated with sNKi gene, compared with that infiltrated with sNK gene. Fibrinolytic activity was detected in transiently expressed samples of two synthetic genes by fibrin plate method, which indicated that the target genes can be normally translated and show thrombolytic activity in tobacco leaves. The translational expression level of sNKi gene was significantly higher than that of sNK gene. Those proved that intron can significantly improve the transient expression of sNK gene.



Key wordsNattokinase      Codon optimization      Transient expression      Intron      Tobacco     
Received: 13 April 2015      Published: 25 September 2015
ZTFLH:  Q786  
Cite this article:

HAN Lan, JU Lin-fang, NIU Yi-ding, WANG Ying-chun, HASI Agula. Optimization, Synthesis and Transient Expression of Nattokinase Gene in Tobacco (Nicotiana tabacum L.) Leaves. China Biotechnology, 2015, 35(9): 14-20.

URL:

https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20150903     OR     https://manu60.magtech.com.cn/biotech/Y2015/V35/I9/14


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