Development of patatin knockdown potato tubers using RNA interference (RNAi) technology, for the production of human-therapeutic glycoproteins

Abstract Background Patatins encoded by a multi-gene family are one of the major storage glycoproteins in potato tubers. Potato tubers have recently emerged as bioreactors for the production of human therapeutic glycoproteins (vaccines). Increasing the yield of recombinant proteins, targeting the produced proteins to specific cellular compartments, and diminishing expensive protein purification steps are important research goals in plant biotechnology. In the present study, potato patatins were eliminated almost completely via RNA interference (RNAi) technology to develop potato tubers as a more efficient protein expression system. The gene silencing effect of patatins in the transgenic potato plants was examined at individual isoform levels. Results Based upon the sequence similarity within the multi-gene family of patatins, a highly conserved target sequence (635 nts) of patatin gene pat3-k1 [GenBank accession no. DQ114421] in potato plants (Solanum tuberosum L.) was amplified for the construction of a patatin-specific hairpin RNAi (hpRNAi) vector. The CaMV 35S promoter-driven patatin hpRNAi vector was transformed into the potato cultivar Desiree by Agrobacterium-mediated transformation. Ten transgenic potato lines bearing patatin hpRNA were generated. The effects of RNA interference were characterized at both the protein and mRNA levels using 1D and 2D SDS/PAGE and quantitative real-time RT-PCR analysis. Dependent upon the patatin hpRNAi line, patatins decreased by approximately 99% at both the protein and mRNA levels. However, the phenotype (e.g. the number and size of potato tuber, average tuber weight, growth pattern, etc.) of hpRNAi lines was not distinguishable from wild-type potato plants under both in vitro and ex vitro growth conditions. During glycoprotein purification, patatin-knockdown potato tubers allowed rapid purification of other potato glycoproteins with less contamination of patatins. Conclusion Patatin-specific hpRNAi effectively suppressed the expression of a majority of patatin variants in potato tubers via the specific degradation of individual mRNAs of the patatin multi-gene family. More importantly, patatin-knockdown potato tubers appear to be an ideal host for the production of human therapeutic glycoproteins, because they eventually allow fast, easy purification of recombinant proteins, with less contamination from potato glycoprotein patatins.</p

Development of plasma sources and diagnostics for the simulation of fusion edge plasmas

© 2022, The Korean Physical Society.Although the research on divertors and scrape-off layers (SOLs) has been not as focused on as the recent success of the Korean fusion program, a few linear plasma devices have been developed for simulating divertor and SOL plasmas: (1) diversified plasma simulator (DiPS), a versatile linear machine, has been developed for simulations of divertor and space plasmas with various electric probes, such as single, triple, and Mach Probes and gridded energy analyzer. DiPS consists of two major parts: a divertor plasma simulator with a LaB 6 DC plasma source and a space plasma simulator with a helicon RF plasma source, (2) divertor plasma simulator-1 (DiPS-1) is a part of DiPS with only a LaB 6 cathode, where a low-power laser-induced fluorescence (LIF) is added and more electric probe diagnostics are augmented; it is dedicated only for fusion edge and divertor plasmas, (3) Divertor Plasma Simulator-2 (DiPS-2) has been modified from the DiPS-1 by adding a magnetic nozzle with a limiter structure and by removing the helicon source and space chamber. DiPS-2 is a linear plasma device with a 4-inch LaB 6 cathode, the same as DiPS-1, and it is focused on the development of various diagnostics, such as those used for LIF and laser Thomson scattering (LTS) along with various electric probes, on the divertor and scrape-off plasmas and on the plasma-material interaction (PMI) research, such as that of tungsten and graphite as plasma-facing components (PFCs), (4) A Multi-Purpose Plasma (MP 2) device is a renovation of the Hanbit mirror device [Kwon et al., Nucl. Fusion 43, 686 (2003)] with the installation of two plasma sources: LaB 6 (DC) and helicon (RF) plasma sources. A honeycomb-like large-area LaB 6 (HLA-LaB 6) cathode has been developed for the divertor plasma simulation to improve the resistance against the thermal shock fragility for large (8-inch) and high density plasma generation, (5) DiPS-2 has been augmented by adding another cylindrical device, called the Dust interaction with Surfaces Chamber (DiSC) for the generation and diagnostics of dusts. This combined system (DiPS-2+DiSC) has added two more diagnostics: Laser Photo-Detachment (LPD) for dust density and laser Mie Scattering (LMS) for dust size. Moreover, dusts or negative ions have been analyzed by using electric probes and capacitive diagram gauges in Transport and Removal of Dusts (TReD) device.N

Simulations of fusion edge plasmas by linear plasma devices: physics and plasma–material interactions

© 2022, The Korean Physical Society.Because a fusion edge plasma contains various atomic and molecular processes, along with various plasma–material interactions (PMIs) for post-mortem analyses, a linear plasma device can simulate divertor and scrape-off layer (SOL) plasmas with DC edge relevant parameters, although it cannot generate a high ion temperature and toroidicity with much less power density compared to toroidal devices. The Divertor Plasma Simulator-2 (DiPS-2), a linear device with an LaB6 DC cathode, has been used for a few fusion-relevant physics experiments, including edge localized mode (ELM) simulation and edge transport of diffusion and convection. An ELM simulation has been performed by modulating the magnetic field relevant to the pressure modulation of a toroidal device, and the diffusion coefficients of free and bound presheaths have been measured in simulations of divertor or limiter transport. Moreover, the convection of the filament or the bubble expansion to the first wall has also been analyzed. In addition to various atomic and molecular processes in SOL and divertor plasmas, PMIs must be analyzed both on and beneath the surface of the plasma-facing components (PFCs) because of surface modification. Using DiPS-2 and other linear devices along with Korea Superconducting Tokamak Advanced Research (KSTAR), PMIs have been analyzed in terms of the following elements or processes: (1) boronizations, both for dust interactions with the surface chamber (DiSC) and KSTAR device, are analyzed; (2) carbon damage by the dense heat flux of DiPS-2 is experimentally investigated; (3) the density profile of the lithium injection gettering of hydrogen and its transport experiments (LIGHT-1) device is analytically calculated; (4) the effect of nitrogen on the relaxation of the heat flux to the divertor tile is experimentally analyzed; and (5) tungsten as the divertor tile material is analyzed via laser ELM simulations in terms of dust generation and surface modification.N

Construction and Commissioning of PAL-XFEL Facility

The construction of Pohang Accelerator Laboratory X-ray Free-Electron Laser (PAL-XFEL), a 0.1-nm hard X-ray free-electron laser (FEL) facility based on a 10-GeV S-band linear accelerator (LINAC), is achieved in Pohang, Korea by the end of 2016. The construction of the 1.11 km-long building was completed by the end of 2014, and the installation of the 10-GeV LINAC and undulators started in January 2015. The installation of the 10-GeV LINAC, together with the undulators and beamlines, was completed by the end of 2015. The commissioning began in April 2016, and the first lasing of the hard X-ray FEL line was achieved on 14 June 2016. The progress of the PAL-XFEL construction and its commission are reported here.11Nsciescopu

Hard X-ray free-electron laser with femtosecond-scale timing jitter

The hard X-ray free-electron laser at the Pohang Accelerator Laboratory (PAL-XFEL) in the Republic of Korea achieved saturation of a 0.144 nm free-electron laser beam on 27 November 2016, making it the third hard X-ray free-electron laser in the world, following the demonstrations of the Linac Coherent Light Source (LCLS) and the SPring-8 Angstrom Compact Free Electron Laser (SACLA). The use of electron-beam-based alignment incorporating undulator radiation spectrum analysis has allowed reliable operation of PAL-XFEL with unprecedented temporal stability and dispersion-free orbits. In particular, a timing jitter of just 20 fs for the free-electron laser photon beam is consistently achieved due to the use of a state-of-the-art design of the electron linear accelerator and electron-beam-based alignment. The low timing jitter of the electron beam makes it possible to observe Bi(111) phonon dynamics without the need for timing-jitter correction, indicating that PAL-XFEL will be an extremely useful tool for hard X-ray time-resolved experiments.1143Nsciescopu