석유기반 플라스틱의 대체제인 폴리하드록시부틸레이트(polyhydroxybutyrate, PHB)의 기존 추출방법은 분자량 감 소 및 물성 변형을 일으킨다. 본 연구에서는 기능화 된 탄소나노튜브(carbon nanotube, CNT)를 부착한 돌기형 탄소나노튜브 분리막의 여과를 통해 물리적 파쇄를 발생시켜 미생물 내 축적된 PHB를 추출하고자 하였다. 돌기형 탄소나노튜브 분리막의 물리적 파쇄를 확인하기 위해 대장균 용액으로 여과 실험을 수행하여 불활성화를 관찰하였다. 또한 PHB를 축적한 미생물 용 액의 여과를 수행하여 PHB가 추출되었는지 확인하였더니 가장 대표적인 추출방법인 chloroform과 비교하여도 여과로 인한 추출이 4% 높은 성능을 가진 것을 관찰하였다. 본 결과를 통해 친환경적 바이오 플라스틱 회수를 위한 돌기형 탄소나노튜브 분리막의 적용 가능성을 확인하였다.

There are many other detection methods for Bursaphelenchus xylophilus. However, Baermann funnel method, PCR-based methods, which are laborious and time consuming processes that are unsuitable to rapid diagnose the Pine Wilt Disease (PWD) on the field. For these reasons, the aim of our experiment is not only to apply field diagnostic for pine wood nematode (PWN) but also to reduce total time for detection PWN in the pine trees by loop-mediated isothermal amplification (LAMP) with phosphate (Pi)-induced coloration reaction which could be yes or no answer for detection of PWN has not required UV detector but it just could be discriminated by naked eyes within 30 min. Genomic DNA was directly extracted from pine wood chips by Wood Chips Direct Lysis procedure which can be used for LAMP only after simple dilution. Our results suggest that LAMP-Pi detection method, simple and rapid method for detection of PWN, could be applied to the field diagnostic for PWD.

It is difficult to identification between Bursaphelenchus spp. and Pine Wood Nematode (PWN) by morphological characteristics without expertise about nematode taxonomy. Furthermore, Baermann funnel method, which is nematode extraction method from wood chips or soil, requires at least 24 hours to extract nematode that is unsuitable to rapid diagnose the Pine Wilt Disease (PWD). For these reasons, the aim of this experiment is not only to improve accuracy of a PCR based method but also to reduce total experiment time for detection Bursaphelenchus spp. and PWN in the wood chips of PWD infected pine tree. In this experiment, we had been employed two PCR primer sets, which were originated from PWN specific Internal Transcribed Spacer (ITS) sequence region and Bursaphenchus spp. universal mitochondrial Cytocrome Oxidase subunit I (mtCOI) sequence region in order to discrimination between Bursaphelenchus spp. and PWN at the same PCR reaction. This experimental procedure was able to reduce experiment time and cost as well as to improve accuracy of detection than previous PCR based detecting method by not using Baermann funnel method and commercial genomic DNA extraction kit but using direct pine wood chips lysis method.

Sodium-potassium-chloride co-transporter (NKCC) is a membrane bound channel protein that plays a prominent role in a variety of epithelial absorptive, secretory processes and a direct role in cell volume regulation, in which NKCC transports sodium, potassium, and chloride ions across the cell membrane. It has been known that prostaglandin E2 (PGE2) induces an acute cell lysis of specific hemocyte type, oenocytoid, to release prophenoloxidase into the plasma and ouabain (a specific sodium pump inhibitor) inhibits the oenocytoid cell lysis resulting in preventing phenoloxidase activation. However, it is not clear how the intracellular signaling pathway leads to oenocytoid cell lysis in response to PGE2. This study was designed to analyze functional role of NKCC in the cell lysis to release prophenoloxidase. A gene structure of NKCC was derived from cDNA library of Spodoptera exigua hemocyte, NKCC was expressed in all developmental stages and tissues. A real time quantitative RT-PCR showed that bacterial challenge significantly induced its expression. Specific inhibitors of NKCC, bumetanide and chlorothiazide, clearly prevented the cell lysis in a dose dependent manner. When RNA interference using double stranded RNA (dsRNA) specific to NKCC suppressed its expression, the oenocytoid lysis and PO activation was significantly inhibited in response to PGE2. It also reduced nodule formation to bacterial challenge. These results indicate that NKCC is associated with oenocytoid cell lysis probably by increasing cell volume through inward transport of ions in response to PGE2.