Inheritance, Linkage and Possible Use of Fractured Starch Mutant in Barley (Hordeum Vulgare L.)

Kwon, Moosik,Chung, Tae Young 성균관대학교 생명공학연구소 2001 生命工學硏究 Vol.7 No.1

In order to breed barley lines with reduced viscosity of wort, a fractured starch mutant of naked barley cultivar, Nubet, was obtained from the M_2 seeds mutated by the diethyl sulfate treatment. Seeds of this fractured starch mutant were opaque and the endosperm consists of angular, irregular and fractured starch. The mutant was caused by single recessive mutation and assigned by the symbol fra. The gene was located on chromosome 4, distal in long arm by linkage recombinations using translocation homozygote lethal test set. The linkage value between the fractured starch mutant and T2-4a, T2-4d were 26.0±4.9, 34.2±3.1 percent respectively. In addition to the reduced seed size, fewer kernels per spike and higher tillering ability, lower β-glucan viscosity and higher lysine content of the grain were associated with this mutant. β-glucan viscosity of the Nubet grains increased from 3 weeks after anthesis to matury and most of the viscose substances appeared to be stored in the middle of the endosperm tissue. Since the mutant grains showed better milling property as compared to Nubet, it can be used as breeding resources to develope new barley cultivars for malting and milling purpose.

Outer Membrane Protein H for Protective Immunity Against Pasteurella multocida

이정민,Young Bong Kim,Moosik Kwon,Moosik Kwon 한국미생물학회 2007 The journal of microbiology Vol.45 No.2

Pasteurella multocida, a Gram-negative facultative anaerobic bacterium, is a causative animal pathogen in porcine atrophic rhinitis and avian fowl cholera. For the development of recombinant subunit vaccine against P. multocida, we cloned and analyzed the gene for outer membrane protein H (ompH) from a native strain of Pasteurella multocida in Korea. The OmpH had significant similarity in both primary and secondary structure with those of other serotypes. The full-length, and three short fragments of ompH were expressed in E. coli and the recombinant OmpH proteins were purified, respectively. The recombinant OmpH proteins were antigenic and detectable with antisera produced by either immunization of commercial vaccine for respiratory disease or formalin-killed cell. Antibodies raised against the full-length OmpH provided strong protection against P. multocida, however, three short fragments of recombinant OmpHs, respectively, showed slightly lower protection in mice challenge. The recombinant OmpH might be a useful vaccine candidate antigen for P. multocida.

Bacillus 殺蟲 蛋白質(CryIC)抗體 生産과 그 應用에 관한 硏究 : Its Application To Plant Biotechnology

권무식 성균관대학교 생명과학자원연구소 1997 生命資源科學硏究 Vol.4 No.2

Bacillus thuringiensis spps., gram-positive soil bacteria, are characterized by their ability to produce crystalline inclusions during sporulation. Inclusions are proteins exhibiting a highly specific entomocidal activity. One of the inclusions, CryIC is specifically toxic to lepidopteran insects. The gene CryIC(7.43kb) has been isolated from Bacillus thuringiensis entomocidus 60.5 and ligated to pTZ vector. The recombinant plasmid named to pSB607. The pSB607 was transformed to E.coli JM109. A number of transformants were selected. The CryIC was purified using differential solubility. The protein was treated with trypsin to increase it's toxicity. They were observed by SDS-PAGE. The activated CryIC was used as an immunogen. Some 600㎍ of the immunogen were hypodermally injected on the hide back & thighs of a rabbit to generate antisera against CryIC. Titration of anti-CryIC antisera has been achieved by blotting or ELISA. In western blot, 2.34ng antigen was detected. A mutant clone of the CrylC is under construction.

Klebsiella pneumonia에서 Quinoprotein의 발현에 대한 면역학적 고찰

권무식 성균관대학교 생명공학연구소 2000 生命工學硏究 Vol.6 No.1

Pyrroloquinoline quinone (PQQ) is an molecule found in many enzymes called quinoproteins, i.e., alcohol dehydrogenases, amine oxidases, decarboxylases, in a broad spectrum of biological materials. It has been considered a very important molecule for physiological property as a enzymatic cofactor. It has been, therefore, interesting to develop a sensitive method to identify the quinoproteins in organisms. Monospecific polyclonal antiserum specific to PQQ was raised in a rabbit as fllow. The PQQ was conjugated to bovine serum albumin (BSA) by 1-Ethyl-3-(3-dimethyl-aminopropyl) carbodiiminde HCI (EDC) mediated reaction. The PQQ-BSA conjugate was purified by gel filtration. The conjugant was emulsified with adjuvant. The emulsion was hyperdermaly injected to a rabbit. PQQ-lipase conjugate was made to use as an antigen for the anti PQQ-BSA conjugate antiserum. The titer of the antiserum was determined by indirect solid phase immunoassay. The antiserum showing the highest titer recognised nano-gram quantity of PQQ-lipase conjugate. Also, we raised anti PQQ antiserum in a rabbit to examine quinoproteins in a prokaryote, Klebsiella, pneumonia.

雜種 自植系統을 利用한 벼의 RELP 遺傳子 地圖 作成

권오병,이정민,권무식 성균관대학교 생명과학자원연구소 1996 生命資源科學硏究 Vol.2 No.2

Numerous genetic studies have been focused on the elucidation of the structure of rice genome because of its economic importance in the world as well as its shortest genome size among monocots. Restriction fragment length polymorphism(RFLP) is a leading method to map a genome. It has been proven that recombinant inbred lines(RILs) provide an efficient way of mapping genetic markers. In this study, RFLPs of RILs of the rice were determined as follows. Some 164 F_11 individuals of RILs have been derived from F_2 populations of Milyang23 × Gihobyeo. Genomic DNAs were isolated from both the parents and RIL individuals. The DNAs were restricted by 8 different endonucleases (BamH I , Dra I , EcoR I , HindⅢ, EcoRⅤ, Xba I , Sca I , and Kpn I ). The restriction fragments were resolved on agarose gel(0.9%) for Southern blots. RFLP patterns of the parents and RIL individuals were determined using the cDNA clones of RZ & KCD, and the genomic clones of Pst I-digested RG as probes. The RFLP patterns and the linkage map of the genome were determined with the aid of a computer program, Macintosh Mapmaker 3.0. The map shows 125 genetic markers including 106 RFLP loci(4 KCD, 67 RG, and 35 RZ clones). This molecular map contains 1,196.8 centi-Morgan(cM) with average interval size of 9.57 cM on the framework map. This study could facilitate chromosome walking, map-based cloning, quantitative trait mapping, marker-assisted breeding, and evolutionary studies of the rice.

A Gene Mutation of Human Dihydrolipoamide Dehydrogenase cDNA

Ryou, Chongsuk,Kwon, Moosik 성균관대학교 생명공학연구소 1999 生命工學硏究 Vol.5 No.1

Mammalian pyruvate dehydrogenase complex (PDC) catalyzes oxidative decarboxylation of pyruvate to produce Co2, acetyl-CoA, and NADH. Dihydrolipoamide Dehydrogenase (E3) is a component of the complex, and the enzymatic deficiency leads to lactic acidosis, increased concentrations of branched-chain amino acids in the plasma and increased urinary excretion of alpha-keto acids. The E3 deficiency also causes neurological degeneration due to the sensitivity of the central nervous system to defects in oxidative metabolism. In this study, an E3 mutant cDNA was generated from a patient showing some pyruvate metabolic defect. The mutant cDNA was subcloned into pBluescript SK-, and nested deletion set of the clone were prepared for the analysis of the whole nucleotide sequence. A silent mutation found in the clone meaning that there was no changes in amino acid sequence.

배추(Brassica campestris L. var. pekinensis Makino)의 cDNA library 구축 및 상동성 비교

안주미,권오병,전봉균,이풍연,이정민,권무식 성균관대학교 생명과학자원연구소 1995 生命資源科學硏究 Vol.2 No.1

Expressed sequence tag(EST) has a good value to discover a new gene or to study its structure. Some 20 ESTs were generated to obtain new genetic resources of chinese cabbage(Brassian campestris L. var. pekinensis Makino). Poly A+ RNAs were isolated from 10-day-old seedlings grown at 25℃ under the day light. cDNA gene bank was constructed using λ ZAP /cDNA synthesis /Giga Pack Gold Packing kit. About a million clones were able to obtain from the library. All the clones examined so far had insert DNAs. Nucleotide compositions of randomly selected clones were determined by the Sanger mthod. The DNA sequences were compared with those deposited in the GenePept and GenBank database to figure out nucleotide homologies. Two ESTs showed significant similarities to the enlisted sequences. They are chloroplast GADPH subunit of Arabidopsis thaliana and carbonic anhydrase of A. thaliana. The full DNA sequences of the two clones are being determined. The cDNA gene bank constructed. in this experiment will being used to isolate more genes induced by the light in the plant.

韓國型 Turnip Mosaic Virus의 純化

김병훈,권오병,마병철,강미란,권무식 성균관대학교 생명과학자원연구소 1996 生命資源科學硏究 Vol.3 No.1

Chinese cabbage(Brassica campestris subsp pekinensis) useful for making a traditional Korean food, Kimchi, has been considered as the most important vegetable in Korea. In order to supply large quantity of chinese cabbage to the consumer, the mass cultivation is demanded. However, the mass production has been faced to serious problem due to fungal, bacterial and/or viral infections. It has been reported that the viral infections are the worst phatogeruc causes in a number of plants. In the chinese cabbage, turnip mosaic virus(TuMV) is the most serious pathogen. Five different strains of TuMV(Cl, C2, C3, C4, and C5) were identified in the world. We are reporting two strains(C4, C5) purified from Brassica nigra. They are all filamentous on electron micrographs. The TuMV C4 is about 1,300nm long, whereas the C5 is about 1,200nm long in average. They are larger than those reported previously(680-900nm). They will be used to produce monoclonal antibodies against the TuMV coat proteins. The antibodies arc required to develop a sensitive and specific screening methods for the TuMV infected plants.

한국인에서 혈액응고인자 Ⅷ 유전자의 제한효소 절편길이 다형성에 관한 연구

전봉균,이풍연,권무식 성균관대학교 생명과학자원연구소 1994 生命資源科學硏究 Vol.1 No.2

Hemophilia A, an X chromosome-linked bleeding disorder affecting 1 in 5,000 males worldwide, due to the defect of blood coagulation factor Ⅷ. The wide range of clinical severity exhibited by haemophiliacs plus the hight incidence of sporadic cases suggest that hemophilia A be caused heterogeneous mutation of the gene. Restriction fragment length polymorphism(RFLP) is being used for prenatal diagnosis or carrier detection of the genetic disorder, However, the polymorphisms exhibits heterogeniety among races indicating that data from a people may not be applied to others. Two polymorphic sites(Bcl Ⅰ and Hind Ⅲ) of the factor Ⅷ gene were examined in Korean employing polymerase chain reaction(PCR). The PCR product of Bcl Ⅰ restriction was 948 nucleotides(nts) long, while that of Hind Ⅲ was 730 nts long. The former(Bcl Ⅰ) can generate three fragments(90nts, 100nts, and 480nts from 5' to 3' direction) with the polymorphic site, or two fragments(190nts and 480nts from 5' to 3' direction) wihout the polymorphic site. The latter(Hind Ⅲ) can generate three fragments(148nts, 286nts, and 514nts from 5' to 3' direction) with the polymorphic site, or two fragments(434nts and 514nts from 5' to 3' direction) without the polymorphic site. The heterozygote frequency calculated from the allele frequencies(0.754/0.246) of the Bcl Ⅰ (intron 18) polymorphism was 37.1%. The heterozygote frequency calculated from the allele frequencies(0.807/0.193) of the Hind Ⅲ polymorphism(intron 19) was 31.2%. Thus, the two intragenic polymorphisms predicted to be informative was 56.7% in these studies.

Pyrroloquinoline quinone(PQQ)에 特異的인 抗體의 生産

마병철,유형진,권무식 성균관대학교 생명과학자원연구소 1997 生命資源科學硏究 Vol.4 No.1

There exist biocatalysts(e.g., some oxidoreductase and dehydrogenase) containing pyrroloquinoline quinone (PQQ) as a cofactor. They are physiologically and pathogenically very important, since the quinoproteins are involved in various cellular activities, such as stimulation of cell growth, pollen germination, protective effects against hepatotoxin-induced liver injury, and so on. It is, therefor, interesting to develop a sensitive method to identify the quinoproteins in organisms. Monospecific polyclonal antiserum specific to PQQ was raised in a rabbit as follow. The PQQ was conjugated to bovine serum albumin (BSA) by 1-Ethyl-3-(3-dimethyl-aminopropyl)carbodiiminde HCl (EDC) mediated reaction. The PQQ-BSA conjugate was purified by gel filtration. The conjugate concentration was measured by Bradford method. The conjugates was emulsified with adjuvant. The emulsion was hyperdermally injected to a rabbit. PQQ-lipase conjugate was made to use as an antigen for the anti PQQ-BSA conjugate antiserum. The titre of the antiserum was determined by indirect solid phase immunoassay. The antiserum showing the highest titre recognised nano-gram quantity of PQQ-lipase conjugate.