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Development of TaqMan probe-based real-time PCR for rapid identification of beef, pork and poultry meat (소, 돼지, 가금육류의 신속한 동정을 위한 TaqMan probe를 이용한 real-time PCR 개발)

  • Koh, Ba-Ra-Da;Kim, Ji-Yeon;Na, Ho-Myung;Park, Seong-Do;Kim, Yong-Hwan
    • Korean Journal of Veterinary Service
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    • v.35 no.3
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    • pp.215-222
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    • 2012
  • Species-specific $TaqMan^{(R)}$ probe-based real-time PCR assays were developed for detection of beef, pork, chicken, duck, goose and turkey. The primer and probe sets used in this study were designed to be complementary to fibroblast growth factor (FGF) for cattle and pig, mitochondrial NADH dehydrogenase (ND) subunit 3 and ND2 for chicken and duck, 12S rRNA for goose and turkey, respectively. As internal positive control we used conserved region in the ribosomal 18S RNA gene to ensure the accuracy of the detection of target DNA by real-time PCR. We confirmed that real-time PCR assays with the primer and probe sets were positive for cattle, pig and chicken intended target animal species with no cross-reactivity with other non-target animal species. Only >50 ng DNA of beef show cross-reactivity in the determination of duck. Using species-specific primer and probe sets, it was possible to detect amounts of 0.1 ng DNA of cattle and pig, 1.0 pg DNA of chicken, duck and turkey, and 0.1 pg DNA of goose for raw samples, respectively. The detection limits were 0.1 ng DNA of cattle, 1.0 ng DNA of pig and 1.0 pg DNA of chicken for DNA mixtures (beef, pork and chicken) extracted from heat-treated ($121^{\circ}C$/5 min) meat samples. In conclusion, it can be suggested that the $TaqMan^{(R)}$ probe-based assay developed in this study might be a rapid and specific method for the identification of meat species in raw or cooked meat products.

Background $K^+$ channel currents in WEHI-231 cells, immature B lymphocytes

  • Nam, Joo-Hyun;Woo, Ji-Eun;Kim, Tae-Jin;Uhm, Dae-Yong;Kim, Sung-Joon
    • Proceedings of the Korean Biophysical Society Conference
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    • 2003.06a
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    • pp.39-39
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    • 2003
  • In our previous study, WEHI-231, an immature B cell line, showed intractable increase in [C $a^{2+}$]$_{c}$ after the B-cell receptor (BCR) ligation and treatment with 2-aminoethoxydiphenylborate (2-APB), which was never observed in Bal-17, a mature B cell line (Nam et al., 2003, FEBS Lett). In this study, a whole cell voltage clamp study revealed a specific expression of a novel type of $K^{+}$ current, namely voltage-independent background-type $K^{+}$ channels (IK-bg), in WEHI-231 cells. IK-bg was dramatically increase by the application of 2-APB (50 $\square$M), which induced severe hyperpolarization of WEHI-231 from -45 ㎷ to -90 ㎷, When dialyzed with $Mg^{2+}$ and ATP-free pipette solution, a spontaneous development of IK-bg and membrane hyperpolarization were observed. IK-bg was insensitive to classical $K^{+}$ channel blockers (TEA, glibenclamide, $Ba^{2+}$(1 mM)), whereas blocked by quinine and quinidine in a voltage-dependent manner ($IC_{50}$/=6~9 $\square$M at +60㎷). Phorbol myrstate, a PKC activator, decreased the amplitude of IK-bg. Extracellular acidification (pH 6.5) slightly inhibited IK-bg. Arachidonic acid, riluzole, or hyposmotic stress could not affect the IK-bg after the full development by the intracellular dialysis with Mg-ATP-free solution. In a cell-attached mode of single channel recording from WEHI231, we found two types of voltage-independent $K^{+}$ channels with unitary conductance of 300 pS and 120 pS, respectively. Both channels showed very short mean open times and their open probabilities were increase by the application of 2-APB. In Bal-17 cells, no such $K^{+}$ current was observed in 50 cells tested. In summary, WEHI-231 immature B cells express background $K^{+}$ channels. The pharmacological properties and the large unitary conductance suggest that novel types of two-pore domain $K^{+}$ channels (2-P-K channels) might be expressed in WEHI-231, which may provide an intriguing targets of signal transduction in the immature B lymphocytes.e B lymphocytes.

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Effents of Plant Explant Position of Miscanthus and Medium Supplements on Callus Induction (억새(Miscanthus spp.) 식물재료와 배지 첨가물질이 캘러스 형성에 미치는 영향)

  • Kim, Kwang-Soo;Kwon, Da-Eun;Lee, Ji-Eun;Cha, Young-Lok;Moon, Youn-Ho;Song, Yeon-Sang;Kang, Yong-Ku
    • Proceedings of the Plant Resources Society of Korea Conference
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    • 2018.04a
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    • pp.54-54
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    • 2018
  • 억새(Miscanthus spp.)는 우리나라 등 동아시아가 원산이며 바이오매스량이 많아 바이오에너지 원료작물로서 잠재성이 크기 때문에 2세대 바이오에탄올 생산 원료로 주목을 받고 있고, 축산깔개 및 토양개량제 등으로도 이용되고 있다. 미국과 유럽 등에서는 생태계 교란 방지를 위해 4배체 물억새(M. sacchariflorus)와 2배체 참억새(M. sinensis)의 종간 교잡 이질 3배체인 불임성 억새(M. x giganteous)을 주로 재배하고 있으나, 단일 유전형의 품종으로 병해충과 자연재해에 취약하여 다양한 억새 품종의 개발이 시급한 실정이다. 본 연구는 억새를 재료로 하여 반수체 및 배수체 확보를 통한 배수성 별 특성 평가와 함께 기내배양을 통해 탈분화 및 재분화 시스템을 구축하여 억새의 육종 효율을 높이기 위해서 실시하였다. 억새 종자로부터 캘러스의 유도는 MS배지와 N6배지에 1mg/L 2.4-D를 첨가하였을 때 비배발생 캘러스(nonembryogenic callus)가 유도되었고, N6배지에 3~5 mg/L 2,4-D를 첨가하였을 때 배발생 캘러스(embryogenic callus)가 발생하였다. MS배지보다는 N6배지에서 캘러스 유도율이 높았으며, 식물생장조절제로 2,4-D와 BA 조합처리 보다 2,4-D 단용 처리하였을 때 캘러스 유도율이 더 높았다. 억새 종에 따른 캘러스 유도율은 물억새가 25.2~49.3%, 참억새는 30.3~52.0%였고 거대억새는 62.6~81.1%로 나타났다. 억새 신초 및 줄기로부터의 캘러스 유도율은 물억새가 4.4~17.2%, 참억새는 1.8~7.7%, 거대억새는 15.3~19.9%로 나타나 종자에 비해 매우 낮았다. 미성숙화기로부터의 캘러스 유도는 억새 종에 따른 차이가 없었으며, 3mg/L 2,4-D 첨가 배지에서 캘러스 유도율이 비교적 낮았고(60~80%), 1mg/L와 5mg/L의 2,4-D가 첨가된 배지에서 캘러스 유도율이 높게(90~95%) 나타났다.

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Development of species-specific multiplex PCR assays of mitochondrial 12S rRNA and 16S rRNA for the identification of animal species (식육감별을 위한 미토콘드리아 12S rRNA와 16S rRNA 유전자의 종 특이적 multiplex PCR 기법 개발)

  • Koh, Ba-Ra-Da;Kim, Ji-Yeon;Na, Ho-Myung;Park, Seong-Do;Kim, Yong-Hwan
    • Korean Journal of Veterinary Service
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    • v.34 no.4
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    • pp.417-428
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    • 2011
  • Species-specific PCR assay was developed for detection of cattle, sheep, goat, horse, dog, pig, chicken, duck, goose, and turkey using mitochondrial 12S rRNA and 16S rRNA as target genes. Also, an internal positive control was used to detect possible false negatives by using 18S rRNA gene. We designed species-specific primers with amplicon length of 190, 219, 350, 467, 241, 119, 171, 229, 111 and 268 bp for cattle, sheep, goat, horse, dog, pig, chicken, duck, goose, and turkey respectively. The specificity of the primers was tested against the other 10 non-target animal species and a cross-reaction was not observed. We developed two multiplex PCR assays for the simultaneous identification of Korea's major livestock species (cattle, pig, chicken and duck) and poultry species (chicken, duck, goose and turkey) from analogous samples, retaining the same specificity. The limit of detection of the multiplex PCR assay (cattle, pig, chicken and duck) ranged between 1 pg and 0.1 pg of template DNA extracts from raw meat. Applying multiplex PCR assays to DNA extracts from experimental pork/beef and pork/chicken tested raw and heat-treated ($120^{\circ}C$ for 30 min) mixtures respectively, detection limit was 0.1% level beef in pork, pork in beef and chicken in pork and 1.0% level pork in chicken. In conclusion, this assay using gel-based capillary electrophoresis would be very useful in highly sensitive and rapid identification of animal species or ingredients in minced meat and other meat products.

Adhesion Performance of UV-curable Debonding Acrylic PSAs with Different Thickness in Thin Si-wafer Manufacture Process (박막 실리콘 웨이퍼용 UV 경화형 Debonding 아크릴 점착제의 두께별 접착 물성)

  • Lee, Seung-Woo;Park, Ji-Won;Lee, Suk-Ho;Lee, Yong-Ju;Bae, Kyung-Rul;Kim, Hyun-Joong;Kim, Kyoung-Mahn;Kim, Hyung-Il;Ryu, Jong-Min
    • Journal of Adhesion and Interface
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    • v.11 no.3
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    • pp.120-125
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    • 2010
  • UV-curable acrylic Pressure-sensitive adhesives (Acrylic PSAs) are used in many different parts in the world. A wafer manufacture process which is based on semiconductor industry is one thing. We have used acrylic PSAs whose thickness is different from $20{\mu}m$ to $30{\mu}m$ in wafer manufacture process so far. But as wafers become more thinner, acrylic PSAs are supposed to satisfy the requirements such as proper adhesion performance. The main purpose of this research is studying proper adhesion performance and UV-curing behavior of UV-curable acrylic PSAs with very thin thickness and then determining optimized conditions to raise the efficiency of thin wafer production. Acrylic PSAs contain 2-Ethylhexyl Acrylate (2-EHA), Acrylic Acid (AA) and Butyl Acrylate (BA). Ethyl acetate (EtAc) is used as solvent. The acrylic PSAs are obtained using solvent polymerization. Thickness of UV-curable acrylic PSAs is different from $10{\sim}30{\mu}m$. By peel strength and probe tack, adhesion performance and UV curing behavior of acrylic PSA are concerned.

Effect of De-graphitization Heat Treatment on Interfacial Bonding Properties of Flake Graphite Cast Iron-Aluminum Dissimilar Materials Produced by High Pressure Die Casting (고압 다이캐스팅법으로 제조한 편상흑연주철 -알루미늄 이종소재의 계면접합특성에 미치는 탈흑연 열처리의 영향)

  • Yang, Ji-Ba-Reum;Kim, TaeHyeong;Jeong, JaeHeon;Kim, SangWoo;Kim, YoonJun;Kim, DongEung;Shin, JeSik
    • Journal of Korea Foundry Society
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    • v.41 no.6
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    • pp.535-542
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    • 2021
  • In this study, to improve the interfacial bond strength of cast iron-aluminum dissimilar materials, graphite was removed to a certain depth from the cast iron surface through de-graphitization heat treatment. As the heat treatment time increased, the depth at which graphite was removed increased, showing a linear relationship between the heat treatment time and depth. Aluminum was filled to a certain depth on the de-graphitized cast iron surface through die-casting method, and no intermetallic compounds were formed on the cast iron-aluminum interface. The interfacial bonding strength showed a value of 90 MPa regardless of the heat treatment time, which is very high compared to the 12MPa bonding strength of the material without de-graphitization heat treatment. This result is thought to be due to the mechanical bonding of the undercut structure as the liquid aluminum, penetrated by the high pressure die-casting process, solidified in the de-graphitized region of the cast iron.

Isolation and molecular characterizations of canine distemper virus from a naturally infected Korean dog using Vero cells expressing dog signaling lymphocyte activation molecule

  • Yang, Dong-Kun;Kim, Ha-Hyun;Lee, Siu;Yoon, Yoon-Seek;Park, Jungwon;Oh, Dongryul;Yoo, Jae Young;Ji, Miryeon;Han, Bokhee;Oh, Subin;Hyun, Bang-Hun
    • Journal of Veterinary Science
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    • v.21 no.5
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    • pp.64.1-64.14
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    • 2020
  • Background: Canine distemper virus (CDV) infection results in high morbidity and mortality in dogs. There has been no report about Isolation of Korean CDV since 1980 in Korea. Objectives: To investigate the biological properties and the genetic characterization of Korean CDV. Methods: Vero cells expressing dog signaling lymphocyte activation molecule (dSLAM) gene named as Vero/dSLAM were used to isolate CDV using 17 samples. Diagnostic methods such as cytopathic effects, immunofluorescence assay, peroxidase linked assay, electron microscopy, rapid immunodiagnostic assay, and reverse transcription polymerase chain reaction were used to confirm the Korean CDV isolate as a CDV. The genetic analysis was performed through cloning and sequencing of hemagglutinin gene of CDV isolate. Results: A virus propagated in Vero/dSLAM cell was confirmed as CDV (CD1901 strain) based on the above methods. The CD1901 strain showed the highest viral titer (105.5 50% tissue culture infectious dose [TCID50]/mL) in the Vero/dSLAM cells at 4 days post inoculation, but did not form a fork on chorioallantoic membrane of 7-day-old egg. Ribavirin, a nucleotide analogue anti-viral agent, inhibits moderately the Korean CDV propagation in the Vero/dSLAM cells. The nucleotide and amino acid sequences of the H gene of CD1901 strain were compared with those of other CDV strains. The CD1901 strain belonged to Asia 1 group and had the highest similarity (99.9%) with the BA134 strain, which was isolated in China in 2008. Conclusions: We constructed successfully Vero/dSLAM and isolated one Korean CDV isolate (CD1901 strain) from a naturally infected dog. The CD1901 strain belonged to Asia 1 genotype.

Trace element analysis of korean car windshield using LA-ICP-MS (LA-ICP-MS를 이용한 한국 자동차 유리의 미량원소 분석)

  • Min, Ji-Sook;Choi, Man-Sik;Heo, Sang-Cheol;Kim, Jae-Kyun
    • Analytical Science and Technology
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    • v.22 no.3
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    • pp.235-246
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    • 2009
  • The analyses of minor and trace elements in glass debris were performed using LA-ICP-MS in order to identify manufacturers using real commercial samples. At first, a calibration curve was made using standard glass samples of NIST 610, 612, 614 and 616. $^{29}Si$ was used as an internal standard, and the ratios of metal/Si for each metal were compared with their concentrations. Based on elements in each sample and standard materials, 24 metals were quantified and the LOD in analysis, according to the blank sample, was in the range of 0.11 mg/kg (Ti)-4.91 mg/kg (Ca). Eleven samples from two manufacturers were collected and five sub-samples were taken from each sample for analysis. 15 elements (Co, Ce, Ca, Mn, Sr, Ba, Li, Rb, U, La, Th, Na, Al, Zr and Hf) were selected to identify manufacturers because some elements (Cu, Cr, Cd and Ni) were below the detection limit and some elements (Ti, Pr, Mg, Nb, Nd) were absent in the analysis of standards and others (Pb and Sn) had a problem of homogeneity. The attempts to identify manufacturers and the manufacturing period were performed through a triangular diagram. In the manufacturer discrimination by discriminant analysis, a canonical discriminant function was made based on Mn, Ce and Rb, and each sample could be identified.

Improvement of Pre-harvest Sprouting Resistance in Korean japonica Varieties through a Precision Marker-based Breeding

  • Kamal Bhattarai;Patricia Izabelle Lopez;Sherry Lou Hechanova;Ji-Ung Jeung;Hyun-Sook Lee;Eok-Keun Ahn;Ung-Jo Hyun;Jong-Hee Lee;So-Myeong Lee;Jose E. Hernandez;Sung-Ryul Kim
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2022.10a
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    • pp.269-269
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    • 2022
  • Pre-harvest sprouting (PHS) on rice panicles is getting problematic in recent several years in Korea due to climate changes such as high temperature and more frequent typhoons during harvesting season. PHS negatively affects grain quality severely and also yield. Genetic improvement of Korean varieties (Oryza sativa ssp. japonica) through a marker assisted-backcross breeding (MAB) with the known PHS resistant genes must be one of ideal solutions. However, the final breeding products of MAB occasionally exhibit unwanted traits, especially the cross between genetically distant parents. This might be caused by linkage drag and/or presence of the gene-unlinked donor introgressions, resulting that the final products could not be released to the farmers. The major PHS resistance gene, Sdr4 (Seed dormancy 4) originated from an indica cultivar, Kasalath was selected as a donor gene. In order to avoid unexpected phenotypes in the breeding products, we performed a precision marker-based breeding (PMBB) consisting of foreground, recombinant, and background selections (FS, RS, and BS) which aim to develop 'single small introgression lines' (~100 kb introgression). Korean varieties (Ilpum and Gopum) were crossed with Kasalath. We developed Sdr4-allele specific markers for FS and a set of polymorphic flanking markers near the Sdr4 (-350kb and +420kb) for RS. To minimize linkage drag, the small introgression (< 125kb) containing Sdr4 was selected in Ilpum background (BC2F4) through 1st RS with ~1,200 F2 or BC1F2 plants (one side trimmed) and then 2nd RS with ~1,000 progenies from the 1st RS selected plants (another side trimmed). After RS, the selected lines were genotyped by using Infinium 7K SNP chip to detect other donor introgressions and the lines were backcrossed. Currently BS is on-going from the backcross-derived progenies with BS markers to remove residual introgressions. During the PMBB process, genetic effect of Sdr-4-Kasalath allele was confirmed in Ilpum and Gopum backgrounds by PHS phenotyping using the segregating BC2F3 or BC1F4 materials. The Sdr4 PMBB lines in Ilpum background (< 125kb introgression) will be valuable genetic resources to improve PHS resistance in modem popular temperate japonica varieties.

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A Study on The 'Kao Zheng Pai'(考證派) of The Traditional Medicine of Japan (일본 '고증파(考證派)' 의학에 관한 연구)

  • Park, Hyun-Kuk;Kim, Ki-Wook
    • Journal of Korean Medical classics
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    • v.20 no.4
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    • pp.211-250
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    • 2007
  • 1. The 'Kao Zheng Pai(考證派) comes from the 'Zhe Zhong Pai' and is a school that is influenced by the confucianism of the Qing dynasty. In Japan Inoue Kinga(井上金娥), Yoshida Koton(吉田篁墩) became central members, and the rise of the methodology of historical research(考證學) influenced the members of the 'Zhe Zhong Pai', and the trend of historical research changed from confucianism to medicine, making a school of medicine based on the study of texts and proving that the classics were right. 2. Based on the function of 'Nei Qu Li '(內驅力) the 'Kao Zheng Pai', in the spirit of 'use confucianism as the base', researched letters, meanings and historical origins. Because they were influenced by the methodology of historical research(考證學) of the Qing era, they valued the evidential research of classic texts, and there was even one branch that did only historical research, the 'Rue Xue Kao Zheng Pai'(儒學考證派). Also, the 'Yi Xue Kao Zheng Pai'(醫學考證派) appeared by the influence of Yoshida Kouton and Kariya Ekisai(狩谷掖齋). 3. In the 'Kao Zheng Pai(考證派)'s theories and views the 'Yi Xue Kao Zheng Pai' did not look at medical scriptures like the "Huang Di Nei Jing"("黃帝內經") and did not do research on 'medical' related areas like acupuncture, the meridian and medicinal herbs. Since they were doctors that used medicine, they naturally were based on 'formulas'(方劑) and since their thoughts were based on the historical ideologies, they valued the "Shang Han Ja Bing Lun" which was revered as the 'ancestor of all formulas'(衆方之祖). 4. The lives of the important doctors of the 'Kao Zheng Pai' Meguro Dotaku(目黑道琢) Yamada Seichin(山田正珍), Yamada Kyoko(山田業廣), Mori Ritsi(森立之) Kitamura Naohara(喜多村直寬) are as follows. 1) Meguro Dotaku(目黑道琢 1739${\sim}$1798) was born of lowly descent but, using his intelligence and knowledge, became a professor as a Shi Jing Yi(市井醫) and as a professor for 34 years at Ji Shou Guan mastered the "Huang Di Nei Jing" after giving over 300 lectures. Since his pupil, Isawara Ken taught the Lan Men Wu Zhe(蘭門五哲) and Shibue Chusai, Mori Ritsi(森立之), Okanishi Gentei(岡西玄亭), Kiyokawa Gendoh(淸川玄道) and Yamada Kyoko(山田業廣), Meguro Dotaku is considered the founder of the 'Yi Xue Kao Zheng Pai'. 2) The family of Yamada Seichin(山田正珍 1749${\sim}$1787) had been medical officials in the Makufu(幕府) and the many books that his ancestors had left were the base of his art. Seichin learned from Shan Ben Bei Shan(山本北山), a 'Zhe Zhong Pai' scholar, and put his efforts into learning, teaching and researching the "Shang Han Lun"("傷寒論"). Living in a time between 'Gu Fang Pai'(古方派) member Nakanishi Goretada(中西惟忠) and 'Kao Zheng Pai' member Taki Motohiro(多紀元簡), he wrote 11 books, 2 of which express his thoughts and research clearly, the "Shang Han Lun Ji Cheng"("傷寒論集成") and "Shang Han Kao"("傷寒考"). His comparison of the 'six meridians'(3 yin, 3 yang) between the "Shang Han Lun" and the "Su Wen Re Lun"("素問 熱論) and his acknowledgement of the need and rationality of the concept of Yin-Yang and Deficient-Replete distinguishes him from the other 'Gu Fang Pai'. Also, his dissertation of the need for the concept doesn't use the theories of latter schools but uses the theory of the "Shang Han Lun" itself. He even researched the historical parts, such as terms like 'Shen Nong Chang Bai Cao'(神農嘗百草) and 'Cheng Qi Tang'(承氣湯) 3) The ancestor of Yamada Kyoko(山田業廣) was a court physician, and learned confucianism from Kao Zheng Pai 's Ashikawa Genan(朝川善庵) and medicine from Isawa Ranken and Taki Motokata(多紀元堅), and the secret to smallpox from Ikeda Keisui(池田京水). He later became a lecturer at the Edo Yi Xue Guan(醫學館) and was invited as the director to the Ji Zhong(濟衆) hospital. He also became the first owner of the Wen Zhi She(溫知社), whose main purpose was the revival of kampo, and launched the monthly magazine Wen Zi Yi Tan(溫知醫談). He also diagnosed and prescribed for the prince Ming Gong(明宮). His works include the "Jing Fang Bian"("經方辨"), "Shang Han Lun Si Ci"("傷寒論釋司"), "Huang Zhao Zhu Jia Zhi Yan Ji Yao"("皇朝諸家治驗集要") and "Shang Han Ja Bing Lun Lei Juan"("傷寒雜病論類纂"). of these, the "Jing Fang Bian"("經方辨") states that the Shi Gao(石膏) used in the "Shang Han Lun" had three meanings-Fa Biao(發表), Qing Re(淸熱), Zi Yin(滋陰)-which were from 'symptoms', and first deducted the effects and then told of the reason. Another book, the "Jiu Zhe Tang Du Shu Ji"("九折堂讀書記") researched and translated the difficult parts of the "Shang Han Lun", "Jin Qui Yao Lue", "Qian Jin Fang"("千金方"), and "Wai Tai Mi Yao"("外臺秘要"). He usually analyzed the 'symptoms' of diseases but the composition, measurement, processing and application of medicine were all in the spectrum of 'analystic research' and 'researching analysis'. 4) The ancestors of Mori Rits(森立之 1807${\sim}$ 1885) were warriors but he became a doctor by the will of his mother, and he learned from Shibue Chosai(澁江抽齋) and Isawaran Ken and later became a pupil of Shou Gu Yi Zhai, a historical research scholar. He then became a lecturer of medical herbs at the Yi Xue Guan, and later participated in the proofreading of "Yi Xin Fang"("醫心方") and with Chosai compiled the "Jing Ji Fang Gu Zhi"("神農本草經"). He visited the Chinese scholar Yang Shou Jing(楊守敬) in 1881 and exchanged books and ideas. Of his works, there are the collections(輯複本) of "Shen Nong Ben Cao Jing"(神農本草經) and "You Xiang Yi Hwa"("遊相醫話") and the records, notes, poems, and diaries such as "Zhi Yuan Man Lu"("枳園漫錄") and "Zhi Yuan Sui Bi"("枳園隨筆") that were not published. His thoughts were that in restoring the "Shen Nong Ben Cao Jing", "the herb to the doctor is like the "Shuo Wen Jie Zi"("說文解字") to the scholar", and he tried to restore the ancient herbal text using knowledge of medicine and investigation(考據). Also with Chosai he compiled the "Jing Ji Fang Gu Zhi"("經籍訪古志") using knowledge of ancient text. Ritzi left works on pure investigation, paid much attention to social problems, and through 12 years of poverty treated all people and animals in all branches of medicine, so he is called a 'half confucianist half doctor'(半儒半醫). 5) Kitamurana Ohira(喜多村直寬 1804${\sim}$1876) learned scriptures and ancient texts from confucian scholar Asaka Gonsai, and learned medicine from his father Huai Yaun(槐園). He became a teacher in the Yi Xue Guan in his middle ages, and to repay his country, he printed 266 volumes of "Yi Fang Lei Ju("醫方類聚") and 1000 volumes of "Tai Ping Yu Lan"("太平禦覽") and devoted it to his country to be spread. His works are about 40 volumes including "Jin Qui Yao Lue Shu Yi" and "Lao Yi Zhi Yan" but most of them are researches on the "Shang Han Za Bing Lun". In his "Shang Han Lun Shu Yi"("傷寒論疏義") he shows the concept of the six meridians through the Yin-Yang, Superficial or internal, cold or hot, deficient or replete state of diseases, but did not match the names with the six meridians of the meridian theory, and this has something in common with the research based on the confucianism of Song(宋儒). In clinical treatment he was positive toward old and new methods and also the experience of civilians, but was negative toward western medicine. 6) The ancestor of the Taki family Tanbano Yasuyori(丹波康賴 912-955) became a Yi Bo Shi(醫博士) by his medical skills and compiled the "Yi Xin Fang"("醫心方"). His first son Tanbano Shigeaki(丹波重明) inherited the Shi Yao Yuan(施藥院) and the third son Tanbano Masatada(丹波雅忠) inherited the Dian You Tou(典藥頭). Masatada's descendents succeeded him for 25 generations until the family name was changed to Jin Bao(金保) and five generations later it was changed again to Duo Ji(多紀). The research scholar Taki Motohiro was in the third generation after the last name was changed to Taki, and his family kept an important part in the line of medical officers in Japan. Taki Motohiro(多紀元簡 1755-1810) was a teacher in the Yi Xue Guan where his father was residing, and became the physician for the general Jia Qi(家齊). He had a short temper and was not good at getting on in the world, and went against the will of the king and was banished from Ao Yi Shi(奧醫師). His most famous works, the "Shang Han Lun Ji Yi" and "Jin Qui Yao Lue Ji Yi" are the work of 20 years of collecting the theories of many schools and discussing, and is one of the most famous books on the "Shang Han Lun" in Japan. "Yi Sheng" is a collection of essays on research. Also there are the "Su Wen Shi"("素問識"), "Ling Shu Shi"("靈樞識"), and the "Guan lu Fang Yao Bu"("觀聚方要補"). Taki Motohiro(多紀元簡)'s position was succeeded by his third son Yuan Yin(元胤 1789-1827), and his works include works of research such as "Nan Jing Shu Jeng"("難經疏證"), "Ti Ya"("體雅"), "Yao Ya"("藥雅"), "Ji Ya"("疾雅"), "Ming Yi Gong An"("名醫公案"), and "Yi Ji Kao"("醫籍考"). The "Yi Ji Kao" is 80 volumes in length and lists about 3000 books on medicine in China before the Qing Dao Guang(道光), and under each title are the origin, number of volumes, state of existence, and, if possible, the preface, Ba Yu(跋語) and biography of the author. The younger sibling of Yuan Yin(元胤 1789-1827), Yuan Jian(元堅 1795-1857) expounded ancient writings at the Yi Xue Guan only after he reached middle age, was chosen for the Ao Yi Shi(奧醫師) and later became a Fa Yan(法眼), Fa Yin(法印) and Yu Chi(樂匙). He left about 15 texts, including "Su Wen Shao Shi"("素間紹識"), "Yi Xin Fang"("醫心方"), published in school, "Za Bing Guang Yao"("雜病廣要"), "Shang Han Guang Yao"(傷寒廣要), and "Zhen Fu Yao Jue"("該腹要訣"). On the Taki family's founding and working of the Yi Xue Guan Yasuka Doumei(失數道明) said they were "the people who took the initiative in Edo era kampo medicine" and evaluated their deeds in the fields of 'research of ancient text', 'the founding of Ji Shou Guan and medical education', 'publication business', 'writing of medical text'. 5. The doctors of the 'Kao Zheng Pai ' based their operations on the Edo Yi Xue Guan, and made groups with people with similar ideas to them, making a relationship 'net'. For example the three families of Duo Ji(多紀), Tang Chuan(湯川) and Xi Duo Cun(喜多村) married and adopted with and from each other and made prefaces and epitaphs for each other. Thus, the Taki family, the state science of the Makufu, the tendency of thinking, one's own interests and glory, one's own knowledge, the need of the society all played a role in the development of kampo medicine in the 18th and 19th century.

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