폐섬유자원의 발효공학적 이용에 관한 연구 (제8보) 섬유소 자화세균의 혼합배양

윤한대,성낙계 ( Han Dae Yun,Nack Kie Sung ) 한국미생물생명공학회 1978 한국미생물·생명공학회지 Vol.6 No.2

규산시용량(珪酸施用量)이 수도(水稻)의 지방산(脂肪酸) 및 Sterol 조성(組成)에 미치는 영향(影響)

양민석,윤한대,허종수,Yang, Min-Suk,Yun, Han-Dae,Heo, Jong-Soo 한국토양비료학회 1983 한국토양비료학회지 Vol.16 No.4

A pot experiment was conducted to find out the effect of application amounts of silicates on the fatty acid and sterol composition of paddy rice at various growing stages. The results obtained were as follows; 1. At maximum tillering stage and heading stage, significant positive correlations were obtained between the ratio of unsaturated fatty acid to saturated fatty acid and silica absorption, dry matter or grain yield, respectively. But significant correlation was not observed between them at harvesting time. 2. At maximum tillering stage and heading stage, the ratio of unsaturated fatty acid to saturated fatty acid in paddy rice was increased with increasing silica application from 80 to 180 ppm of soil, but these ratio was decreased at the plot of 200Kg/10a of silicate. 3. The ratio of sitosterol to stigmasterol in paddy rice at maximum tillering stage was increased with increasing silica application of soil, but these ratio was increased with increasing silica application up to 180 ppm of soil at heading stage, and was decreased at the plot of 200Kg/10a (410 ppm in soil) application of silicates. 1. 식물체(植物體) 중(中) 전생육기간(全生育期間)동안 검출(檢出)된 지방산(脂肪酸)의 종류(種類)는 $C_{12:0}$, $C_{14:0}$, $C_{16:0}$, $C_{16:1}$, $C_{16:2}$, $C_{18:0}$, $C_{18:1}$, $C_{18:2}$, $C_{18:3}$, $C_{20:2}$와 몇종(種)의 미동정(未同定)된 지방산(脂肪酸)으로 구성(構成)되어 있었다. 2. 규산흡수양(珪酸吸收量), 건물중(乾物重) 및 수량(收量)은 unsaturated fatty acid/saturated fatty acid와 최고분얼기(最高分蘖期) 및 출수기(出穗期)에는 유의적(有意的)인 정(正)(+)의 상관(相關)이 있었으며 수확기(收穫期)에는 유의적(有意的)인 상관(相關)은 없었으나 전반적(全般的)으로 역(逆)(-)의 관계(關係)가 있었다. 3. 식물체중(植物體中) UFA/SFA 비(比)의 평균치(平均値)는 최고분얼기(最高分蘖期) 및 출수기(出穗期)에서는 토양중(土壤中) 규산함량(珪酸含量)이 180ppm까지는 규산함량(珪酸含量)이 높을수록 증가(增加)하였고, 규산자원(珪酸資源) 200kg/10a 시비구(施肥區)(약(約) 410ppm)에서는 감소(減少)하였다. 4. 최고분얼기(最高分蘖期)의 sitosterol/stigmasterol 비(比)는 토양중(土壤中) 규산농도(珪酸濃度)가 높을수록 계속 증가(增加하였으나, 출수기(出穗期)에 있어서는 그 비(比)가 토양중(土壤中) 규산농도(珪酸濃度)를 180ppm 조절시(調節時)까지는 증가(增加)하다가 200kg/10a 시용(施用)(410ppm)에서는 감소(減少)하였다.

토양환경 , 미생물 , 영양 : 작물 생육촉진 유용 미생물 분리 , 동정과 고추에 대한 처리효과

이영한,윤한대 ( Young Han Lee,Han Dae Yun ) 한국토양비료학회 1996 한국토양비료학회 학술발표회 Vol.1996 No.56

식물 내생균 Bacillus sp. CY22가 생성하는 iturin isoform의 분리 및 특성

조수정,윤한대,Cho, Soo-Jeong,Yun-Han-Dae Korean Society of Life Science 2005 생명과학회지 Vol.15 No.6

식물 내생균 Bacillus sp. CY22는 식물병원균 Rhizoctonia solani, Fusarium oxysporum 및 Phythium ultimum에 대해 강한 항균력을 나타내었다. 일반적으로 많은 Bacillus속 균주들은 iturin, fengycin, mycosubtulin과 같은 항균 물질을 분비한다. 본 연구에서는 식물내생균 Bacillus sp. CY22의 배양액으로부터 항균물질을 분리, 정제하였으며 MALDI-TOF mass로 분자량을 확인하였다. MALDI-TOF mass spectrum분석 결과 분리된 항균물질은 Bacillus 속 균주가 생성하는 항균물질로서 잘 알려져 있는 iturin의 분자량과 거의 일치하였으며, m/z 1043.4, 1057.4, 1071.4에서 molecular ion peak를 나타내었다. 이들은 각각 m/z 14차이를 가진 iturin의 isoform으로 추정되며 이 것은 iturin을 구성하고 있는 지방산의 탄소수 차이로 생각되며 m/z 1065.4, 1079.4 peak는 sodium adduct로서 추정된다. 또한 항균물질 iturin을 생성하는데 관여하는 transacylase 유전자를 크로닝하여 ita22 유전자로 명명하고, 그 특성으로 ita22 유전자는 400 개의 아미노산을 인지하는 1,200 bp의 open reading frame (ORF)을 가지며, 아미노산의 상동성을 조사한 결과 Bacillus subtilis 168의 FenF (BAB69697)와 가장 유사하였다. Endophytic Bacillus sp. CY22 was previously isolated from the interior of balloon flower root and showed strong antifungal activity against phytopathogenic fungi such as Rhizoctonia solnni, Fusarium oxysporum, and Phythium ultimum. Many Bacillus strains produce antifungal compound such as iturin, fengycin, and mycosubtilin. We isolated and identified antifungal compound from cell supernatant of the endophytic strain. By the MALDI-TOF mass result, the antifungal compound was similar to the known antifungal lipopeptide iturin. It was found that the purified iturin had three isoforms with protonated masses of m/z 1,043.39, 1,057.42, and 1,071.42 and different structures in combination with $Na^{+}$ ion using MALDI-TOF MS. The ita22 gene, which transacylase gene is associated with production of antifungal iturin, had an open reading frame (ORF) of 1,200 bp encoding 400 amino acids. Results of deduced amino acids sequence homology search, Ita22 was homologous with FenF (BAB69697) of Bacillus subtilis 168.

Effect of Soil Physical Characteristics on Rhizome Rot Incidence of Platycodon grangiflorus

Young-Han Lee(이영한),Yong-Jo Choi(최용조),Sang-Ryeol Park(박상렬),Min-Keun Kim(김민근),Soo-Jeong Cho(조수정),Han-Dae Yun(윤한대) 한국토양비료학회 2000 한국토양비료학회지 Vol.33 No.1

Molecular Cloning of Alfalfa(Medicago Sativa L. Vernal) Leghemoglobin Structural Genes Synthesized by $Poly(A)^+$-mRNA from Alfalfa Root Nodules

조무제,윤한대,최용락,최영주,이경상,Cho, Moo-Je,Yun, Han-Dae,Choi, Yong-Lark,Choi, Young-Ju,Lee, Kyung-Sang,Brill, Winston J. 생화학분자생물학회 1985 한국생화학회지 Vol.18 No.1

알팔파 근류에서 분리 정제한 $poly(A)^+$-mRNA로부터 leghemoglobin message 함유비율이 높은 9S, 18S 및 28S mRNA를 이용하여 cDNA를 합성하였다. 이들 cDNA를 Sal I linker에 연결 pBR322의 Sal I site에 크로닝하여 cDNA library를 만든 후 이들 library로부터 colony hybridization, Southern blot hybridization, hybrid-released 및 hybrid-arrestes translation에 의하여 leghemoglobin cDNA clone을 동정하였으며, 동정된 clone으로부터 alfalfa leghemoglobin cDNA의 제한효소 지도를 작성하였다. Complementary DNAs were synthesized by the leghemoglobin message rich 9S, 18S and 28S-mRNA fractions prepared from the $poly(A)^+$-mRNA of alfalfa root nodules. The synthetic Sal I linkered double stranded cDNA was inserted into the Sal I site of the pBR322, and transformed in E. coli HB101. The leghemoglobin cDNA clones were screened by colony hybridization and Southern blot hybridization with the soybean leghemoglobin cDNA as a probe, and further screened by hybrid-released and hybrid-arrested translation. One alfalfa leghemoglobin cDNA clone was selected, and restriction map and fingerprints of the cDNA from the selected clone were analyzed.

Cloning of α-Amylase Gene from Unculturable Bacterium Using Cow Rumen Metagenome

조수정,윤한대,Cho, Soo-Jeong,Yun-Han-Dae Korean Society of Life Science 2005 생명과학회지 Vol.15 No.6

미생물 메타게놈은 특이한 생체촉매의 다양한 원료로 제공된다. 한우의 반추위에서 게놈 DNA를 분리한 후 메타게놈 은행을 구축하고 $\alpha$-amlylase를 암호화하는 유전자를 클로닝하여 DNA 및 아미노산 서열을 밝히고 생화적 특징을 조사하였다. amyA유전자는 1,893 bp로 631개의 아미노산 잔기를 가진 단백질을 암호화였으며 효소의 분자량은 단백질 전기영동 결과 약 71,000 Da으로 확인되었다. 이 효소를 다른 아밀라제와 비교한 결과 $21-59\%$의 상동성을 보였다. AnyA는 pH $6.40\%$에서 최적 활성을 나타내었고, pl값은 5.87이었다. E. coliDH5$\alpha$에서 발현된 AmyA의 활성은 $\Mg^{2+}$(20mM), $Ca^{2+}$ (30 mM) 존재 시 그 활성이 증가하였고, $Fe^{2+}$, $Cu^{2+}$ 존재 시 저해되었다. amyA 유전자의 internal primer를 사용하여 인공적으로 배양할 수 있는 49종의 반추세균에서 분리한 게놈 DNA을 주형으로 PCR분석한 결과 해당하는 벤드를 확인할 수 없었다. AmyA는 현재로 배양할 수 없는 반추 미생물에서 온 것으로 추정된다. The metagenomes of complex microbial communities are rich sources of novel biocatalysts. The gene encoding an extracellular $\alpha$-amylase from a genomic DNA of cow rumen was cloned in Escherichia coli DH5$\alpha$ and sequenced. The $\alpha$-amylase (amyA) gene was 1,893 bp in length, encoding a protein of 631 amino acid residues with calculated molecular weight of 70,734 Da. The molecular weight of the enzyme was estimated to be about 71,000 Da by active staining of a SDS-PACE. The enzyme was 21 to $59\%$ sequence identical with other amyloyltic enzymes. The AmyA was optimally active at pH 6.0 and $40\%$. The AmyA had a calculated pI of 5.87. AmyA expressed in E. coli DH5$\alpha$ was enhanced in the presence of $Mg^{2+}$ (20 mM) and $Ca^{2+}$ (30 mM) and inhibited in the presence of $Fe^{2+}$ and $Cu^{2+}$. The origin of amyA gene could not be confirmed by PCR using internal primer of amyA gene from extracted genomic DNA of 49 species rumen culturable bacteria so far. An amyh is supposed to obtained from unculturable rumen bacterium in cow rumen environment.

알파파 근류로부터 분리한 Poly ( A+ ) - mRNA 에 의하여 합성된 Leghemoglobin 유전자의 cDNA Cloning 에 관한연구

조무제,윤한대,최용락,최영주,이경상,Winstion J . Brill ( Moo Je Cho,Min Suk Yang,Han Dae Yun,Yong Lark Choi,Young Ju Choi,Kyung Sang Lee ) 생화학분자생물학회 1985 BMB Reports Vol.18 No.1

Complementary DNAs were synthesized by the leghemoglobin message rich 9S, 18S and 28S-mRNA fractions prepared from the poly(A)^+-mRNA of alfalfa root nodules. The synthetic Sal I linkered double stranded cDNA was inserted into the SalI site of the pBR322, and transformed in E. coli HB101. The leghemoglobin cDNA clones were screened by colony hybridization and Southern blot hybridization with the soybean leghemoglobin cDNA as a probe, and further screened by hybrid-released and hybridarrested translation. One alfalfa leghemoglobin cDNA clone was selected, and restriction map and fingerprints of the cDNA from the selected clone were analyzed.

염류수분별 고추 생육과 수량에 미치는 植物生育促進微生物 접종효과

이영한(Young-Han Lee),양민석(Min-Suk Yang),윤한대(Han-Dae Yun) 한국토양비료학회 1996 한국토양비료학회지 Vol.29 No.4

Dissimilatory Nitrate Reduction Characteristics of Indigenous Soybean Rhizobia Distributed in Korea Soils

최영주,최용락,윤한대,유진창,이상규,조무제,Choi, Young-Ju,Choi, Yong-Lark,Yun, Han-Dae,Ryu, Jin-Chang,Lee, Sang-Kyu,Cho, Moo-Je 한국응용생명화학회 1986 Applied Biological Chemistry (Appl Biol Chem) Vol.29 No.2

연속적으로 대두를 재배한 토양과 신개간지 토양에서 재배한 대두 품종 단엽의 근류로부터 토착대두 근류근 87종을 분리하고 이들을 생장속도에 따라 Bradyrhizobium japonicum (slow-grower) 및 Rhizobium fredii (fast-grower)로 분류한 결과 전자가 55종, 후자가 32종이었다. 이두그룹의 대두 근류균을 질산염 환원특성에 따라 denitrifying fast-grower(F-1), nitrate respiring fast-grower(F-II), denitrifying slow-grower(S-I), 및 nitrate respiring slow-grower (S-II)로 세분 되었으며, 분리균주 총 87종중 S-I, S-II, F-I 및 F-II의 수는 각각 10, 22, 48 및 7종이었다. 이들 각 group 대두 근류균의 균체단백질의 전기영동 pattern상의의 차이를 1차 및 2차원 전기영동으로 분석한 결과 slow 및 fast-grower간에는 현저한 차이를 관찰할 수 있었으나 동일 생장속도를 가진 것 중 denitrifer와 nitrate repirer간에는 큰 차이가 없었다. Eightyseven strains of indigenous Rhizobia were isolated from the nodule of soybean cultivar, Danyup, cultivated in four different soils sampled from continuously soybean cultivated and newly reclaimed fields. The strains were grouped into Bradyrhizobium japanicum (slow-grower:55 strains) and Rhizobium fredii (fast-grower: 32 strains). The both groups could be divided into two sub-groups according to the denitrification characteristics, that is, denitrifying fast-grower (F-I), nitrate respiring fast-grower (F-II), denitrifying slow-grower (S-I). and nitrate respiring slow-grower (S-II). Among the 87 isolates, F-I, F-II, S-I and S-II sub-groups were 10, 22, 48 and 7 strains, respectively. The one-and two-dimensional polyacrylamide gel electrophoretic pattern of the four sub-groups were compared and discernible difference was observed between fast and slow-grower, but the difference was not discernible between subgroups within the same growth rate group.