• Journal of Microbiology and Biotechnology
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• 제30권9호
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• pp.1430-1435
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• 2020

Bacterial cellulose (BC) has outstanding physical and chemical properties, including high crystallinity, moisture retention, and tensile strength. Currently, the major producer of BC is Komagataeibacter xylinus. However, due to limited tools of expression, this host is difficult to engineer metabolically to improve BC productivity. In this study, a regulated expression system for K. xylinus with synthetic ribosome binding site (RBS) was developed and used to engineer a BC biosynthesis pathway. A synthetic RBS library was constructed using green fluorescent protein (GFP) as a reporter, and three synthetic RBSs (R4, R15, and R6) with different strengths were successfully isolated by fluorescence-activated cell sorting (FACS). Using synthetic RBS, we optimized the expression of three homologous genes responsible for BC production, pgm, galU, and ndp, and thereby greatly increased it under both static and shaking culture conditions. The final titer of BC under static and shaking conditions was 5.28 and 3.67 g/l, respectively. Our findings demonstrate that reinforced metabolic flux towards BC through quantitative gene expression represents a practical strategy for the improvement of BC productivity.

• 한국자기학회지
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• 제11권6호
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• pp.272-277
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• 2001

FeMr에 의해 교환 바이어스된 synthetic antiferromagnet(CoFe/Ru/CoFe)을 가진 Top Ta/NiFe/CoFe/Cu/CoFe/Ru/CoFe/FeMn/Ta 스핀 밸브 구조를 마그네트론 스퍼터링법에 의해 증착하였다. 이러한 스핀 밸브에서는 자유층, 구속층등의 두께 및 조성이 층간 결합력의 세기를 비롯한 자성특성에 영향을 미치게 된다 후방산란법은 두께 및 조성에 대한 절대정량이 가능하며 비파괴 분석법이라는 장점을 지니고 있으나, 원자번호가 20번 이상인 주기율표상의 인접원소로 이루어진 자성박막을 분석하는데 있어서 신호의 중첩현상으로 인해 분석이 불가능하였다 본 연구에서는 element-specific 한 분석기술인 양성자 여기 X선 검출법과, 절대 정량이 가능하고 깊이분해능을 현저히 향상시킨 grazing-exit 후방산란법 (RBS : Rutherford Backscattering Spectrometry)을 동시에 사용하여 상호 보완적인 분석을 함으로써 스핀밸브에 대한 성분 및 두께에 대한 정량분석을 수행하였다. 이를 위하여 먼저 spin valve 구조에서 자성층인 NiFe, CoFe, FeMn 단일층이 증착된 시료에 대한 표준화를 수행함으로써 spin valve 구조에서 grazing-exit 후방산란 스펙트럼 상의 중첩된 신호를 Simulation을 통하여 분리가 가능하였으며, 특히 Ru층의 두께는 단위의 정확도로 측정이 가능 하였다

• Journal of Microbiology and Biotechnology
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• 제15권4호
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• pp.880-886
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• 2005

Using carotenoid genes of Erwinia herbicola, metabolic engineering was carried out for lycopene production with the pAC-LYCO4 plasmid, which was composed of a chromosomal DNA fragment of E. herbicola containing the crtE, crtB, and crtI genes under the control of the tetracycline promoter and the ipi gene of Haematococcus pluvialis with the trc promoter. Plasmid pAC-LYCm4 was constructed for efficient expression of the four exogenous genes using a strong RBS sequence and the same tetracycline promoter. The optimized expression construct of pAC-LYCm4 increased Iycopene production three times as compared with pAC-LYCO4. pAC-LYCm5 containing ispA behind the four exogenous genes was constructed. There was no significant difference in Iycopene production and cell growth between pAC-LYCm4 and pAC-LYCm5. FPP synthase encoded by ispA was not rate-limiting for Iycopene production. Each gene of crtE, crtB, crtI, and ipi was overexpressed, using pBAD-crtE, pBAD-crtIB, and pBAD-ipiHPI, in addition to their expression from pAC-LYCm4. However, there was no increase oflycopene production with the additional overexpression of each exogenous gene. The four exogenous genes appeared to be not rate-limiting in cells harboring pAC-LYCm4. When pDdxs, pBAD24 containing dxs, was introduced into cells harboring lycopene synthetic plasmids, lycopene production of pAC-LYCO4, pAC-LYCm4, and pAC-LYCm5 was increased by 4.7-, 2.2-, and 2.2-fold, respectively. Lycopene production of pBAD-DXm4 containing crtE, crtB, crtI, ipi, and dxs was 5.2 mg/g dry cell weight with $0.2\%$ arabinose, which was 8.7-fold higher than that of the initial strain with pAC-LYC04. Therefore, the present study showed that proper regulation of a metabolically engineered pathway is important for Iycopene production.