• Title/Summary/Keyword: molecular breeding

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Isolation and Characterization of Bacillus subtilis CH-10 Secreting Cellulase from Cattle Manure (우분으로부터 Bacillus subtilis CH-10의 분리 및 균주가 분비하는 Cellulase의 특성에 관한 연구)

  • Kim, Tae-Il;Han, Jung-Dae;Jeon, Byoung-Soo;Ha, Sang-Woo;Yang, Chang-Bum;Kim, Min-Kyun
    • Korean Journal of Microbiology
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    • v.35 no.4
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    • pp.277-282
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    • 1999
  • A bacterium producing the extracellular cellulase was isolated from cattle feces and screened as cellulase activity was excellent upon congo red straining method and activity measurements. Isolate was identified as Bacillus subtilis CH-10 on the basis of morphological and biochemical properties as well as cellular fatty acids composition. The enzyme which the isolate secretes had the optimum initial pH and temperature for its induction was 7.5 and 50${\circ}C$, respectively. The maximum CMCase activity in crude enzyme solution was observed at pH 7.5 and 75${\circ}C$ and was stable for pH 7.5 to 9.0 to maintain 70% activity. When the isolate was cultured in CMC media at 37${\circ}C$ for 24 hrs, CMCase and FPase activity was 1.13 U/㎖and 0.16U/㎖, respectively whereas Avicelase and ${\beta}$-glucosidase activity was not detected. When crude supernatant was used for zymogram, three major bands, cel 1, cel 2 and cel 3, were detected approximately 39, 41 and 57 KDa, respectively on CMC-SDS-PAGE.

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Characteristics of fruiting bodies color mutants in Pleurotus ostreatus (원형느타리버섯 백색돌연변체의 특성)

  • Lee, Kang-Hyo;Kim, Gyu-Hyun;Kim, Beom-Gi;Yoo, Young-Bok;Sung, Jae-Mo
    • Journal of Mushroom
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    • v.5 no.1
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    • pp.34-38
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    • 2007
  • The white-colored and the dark gray-colored mutants were frequently happened in cultivated areas of Pleurotus ostreatus (Wonhyeong-neutari). These caused conflicts between farmers and spawn companies. Our studies were conducted to elucidate the mechanism of mutagenesis. The results from the studies would provide valuable informations that could be used to prevent the color-related mutation, and also will be applied in breeding programs of P. ostreatus. Oyster mushroom variety, Wonhyeong-neutari, is somatic hybrid of Pleurotus and has genetic makers for arginine, ornithine, proline, riboflavine. Genetic markers analysis of monospore isolates derived from color mutants show identical tendency with that of Wonhyeong-neutari, these results indicate that color mutants were derived from Wonhyeongneutari. Twenty-one and four homokaryons were selected from the white-colored mutant MGL 2205 and gray-colored ASI 2029. All 34 F1 hybrids derived from the white-colored mutant MGL 2205 produce white-color fruiting bodies, indicating that the white color trait is heritable. In the first generation hybrids between the white-colored MGL 2205 and the gray-colored ASI 2029, all 16 hybrids produced pigmented fruiting bodies. Homokaryons isolated from the hybrid MGL 2205 X ASI 2029 were mated with homokaryon tester strains derived from MGL 2205. By these result, we could assumed that white color trait is a heritable character which is controlled by more than one recessive gene.

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Efficient Plant Regeneration from Mature Seed-derived Callus of Italian ryegrass (Lolium multiflorum Lam.) (이탈리안 라이그래스의 성숙종자 유래 캘러스로부터 효율적인 식물체 재분화)

  • Woo, Hyun-Sook;Lee, Sang-Hoon;Lee, Dong-Gi;Kim, Jin-Soo;Won, Sung-Hye;Lee, Byung-Hyun
    • Journal of Plant Biotechnology
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    • v.31 no.1
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    • pp.43-48
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    • 2004
  • As an initial step for future genetic manipulations to improve forage characteristics of Italian ryegrass (Lolium multiflorum Lam.), an efficient tissue culture system was established and the factors affecting plant regeneration were evaluated. MS medium containing 5mg/L 2,4-D was optimal for embryogenic callus induction from mature seed and had a strong effect on successive plant regeneration. The plant regeneration frequency was observed at above 70% when embryogenic calli induced were transferred to N6 medium supplemented with 1mg/L 2,4-D and 5mg/L BA. Among several basic media tested, MS and N6 medium were optimal for callus induction and plant regeneration, respectively. Genotype was an important factor in plant regenerability. 'Jeanne' showed the highest regeneration frequency of 73%. A short tissue culture period and high-frequency regeneration system established in this study will be useful for molecular breeding of Italian ryegrass through genetic transformation.

Efficient Screening Method for Resistance of Cucumber Cultivars to Fusarium oxysporum f. sp. cucumerinum (오이 덩굴쪼김병에 대한 효율적인 저항성 검정 방법)

  • Lee, Ji Hyun;Kim, Jin-Cheol;Jang, Kyoung Soo;Choi, Yong Ho;Choi, Gyung Ja
    • Research in Plant Disease
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    • v.20 no.4
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    • pp.245-252
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    • 2014
  • The study was performed to establish an efficient screening method for resistant cucumber to Fusarium oxysporum f. sp. cucumerinum. The isolate KR5 was identified as F. oxysporum f. sp. cucumerinum based on molecular analyses of ITS and TEF genes and host-specificity test on cucurbits including melon, oriental melon, cucumber, and watermelon. Then four cucumber and two rootstock cultivars showing different resistance degrees to the Fusarium wilt pathogen KR5 were selected. And development of Fusarium wilt of the six cultivars according to several conditions, including incubation temperature after inoculation, inoculum concentration, root wounding, and growth stages of seedlings, was investigated. Disease severity of Fusarium wilt on the resistant cultivars was changed with incubation temperatures after inoculation. The resistant cultivars showed the higher resistance when inoculated plants were kept at 25 or $30^{\circ}C$ than at $20^{\circ}C$. Among four different growth stages of the seedlings, seven-day-old seedling represented the most difference of resistance and susceptibility to Fusarium wilt. From above results, we suggest that an efficient screening method for resistant cucumber to F. oxysporum f. sp. cucumerinum is to dip the non-cut roots of seven-day-old seedlings in spore suspension of $1.0{\times}10^6-1.0{\times}10^7$ conidia/ml and to transplant the seedling into a non-infected soil, and then to incubate the inoculated plants in a growth room at $25^{\circ}C$ for 3 weeks to develop Fusarium wilt.

Isolation and Characterization of Pepper mottle virus Infecting Tomato in Korea

  • Kim, Mi-Kyeong;Kwak, Hae-Ryun;Han, Jung-Heon;Ko, Sug-Ju;Lee, Su-Heon;Park, Jin-Woo;Jonson, Miranda Gilda;Kim, Kook-Hyung;Kim, Jeong-Soo;Choi, Hong-Soo;Cha, Byeong-Jin
    • The Plant Pathology Journal
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    • v.24 no.2
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    • pp.152-158
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    • 2008
  • A peculiar virus-like disease of tomato showing yellow mosaic and necrotic spots on leaves and necrosis on veins, petioles and stems was observed at the Tomato Experimental Station (TES), Buyeo, Chungcheongnamdo, Korea. The disease incidence at TES fields ranged from 21 to 35% infecting different tomato cultivars. For this reason, to identify the virus infecting tomato and to characterize the virus based on biology, serology, cytology and at molecular level. Here, leaf samples were randomly collected from different infected tomato cultivars at TES fields and greenhouses and tested by ELISA using Pepper mottle virus (PePMoV) and Tomato mosaic virus (ToMV) antisera. Infected saps were mechanically inoculated in different host plants to test for pathogenicity, symptomatology and host ranges. Infected tissues and ultrathin sections were examined by electron microscopy. Finally, putative coat protein and 3'-untranslated region (CP/3'-UTR) fragment was amplified and cloned for sequence determination and analyzed its genetic relationship to existing PepMoV and PVY sequences at the Genbank. Results showed 69% of the samples were positive with PepMoV, 13% with ToMV and 19 % were doubly infected with PepMoV and ToMV. Symptoms greatly varied from different host plants inoculated with tomato leaf sap infected with PepMoV alone and discussed in detailed in this paper. Electron microscopy from infected tissues showed filamentous particles of 720-750nm in length, a typical morphology and size of PepMoV. In addition, cylindrical inclusion bodies, pinwheels, scrolls and laminates with masses of fibrillar inclusions were also found in ultrathin sections. Alignment of the sequences of the CP/3'-UTR revealed >96% sequence identity with PepMoV and only <61% with PVY. Taken together, all these evidences presented clearly indicated that the causal agent infecting tomato at TES was PepMoV and we designated this PepMoV infecting tomato as Tom-sd2 strain in this study.

Distribution of DArT Markers in a Genetic Linkage Map of Tomato (토마토 유전자연관지도 상의 DarT 마커 분포)

  • Truong, Hai Thi Hong;Graham, Elaine;Esch, Elisabeth;Wang, Jaw-Fen;Hanson, Peter
    • Horticultural Science & Technology
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    • v.28 no.4
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    • pp.664-671
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    • 2010
  • A genetic linkage map was constructed using 188 $F_9$ RILs derived from a cross between $Solanum$ $lycopersicum$ H7996 (resistant to bacterial wilt) and $S.$ $pimpinellifolium$ WVa700 (highly susceptible to bacterial wilt). The map consisted of 361 markers including 260 DArTs, 74 AFLPs, 4 RFLPs, 1 SNP, and 22 SSRs. The resulting linkage map was comprised of 13 linkage groups covering 2042.7 cM. The genetic linkage map had an average map distance between markers of 5.7 cM, with an average DArT marker density of 1/7.9 cM. Based on the distribution of anchor SSR markers, 11 linkage groups were assigned to 10 chromosomes of tomato except chromosomes 5 and 12. The DArT markers were distributed across the genome in a similar way as other markers and showed the highest frequency of clustering (38.8%) at ${\leq}$ 0.5 cM intervals between adjacent markers, which is 3 times higher than AFLPs (13.5%). The present study is the first utilization of DArT markers in tomato linkage map construction.

Effect of Plant Growth Regulators and Medium Supplements on Plant Regeneration of Kentucky Bluegrass (식물생장조절물질과 배지첨가물질이 켄터키 블루그래스의 식물체 재분화에 미치는 영향)

  • Lee Sang-Hoon;Lee Ki-Won;Kim Do-Hyun;Lee Dong-Gi;Won Sung-Hye;Kim Ki-Yong;Lee Byung-Hyun
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.26 no.2
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    • pp.69-76
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    • 2006
  • To optimize tissue culture responses for genetic transformation of Kentucky bluegrass, the effects of culture medium supplements on tissue culture responses were investigated with mature seeds of a cultivar 'Newport' as explant tissues. The optimal concentration of 2,4-D (2.4-dichloro phenoxy acetic acid) for the induction of embryogenic callus from mature seed was 3 mg/L. Plant regeneration frequency was 54% when embryogenic callus was cultured on the regeneration medium supplemented with 1 mg/L 2,4-D and 3 mg/L of BA (6-benzyladenine). Addition of 1 g/L of casein hydrolysate and 500 mg/L of L-proline improved frequencies of embryogenic callus induction and plant regeneration up to 60.8% and 58.3%, respectively. Regenerated plants were grown normally when shoots transplanted to the soil. A rapid and efficient plant regeneration system established in this study. We suggest that the results may be useful for molecular breeding of Kentucky bluegrass through genetic transformation.

Effect of Callus Type and Antioxidants on Plant Regeneration and Transformation of Tall Fescue (캘러스의 형태와 항산화물질 첨가가 톨 페스큐의 식물체 재분화와 형질전환효율에 미치는 영향)

  • Lee Ki-Won;Lee Sang-Hoon;Kim Do-Hyun;Lee Dong-Gi;Won Sung-Hye;Lee Hyo-Shin;Lee Byung-Hyun
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.26 no.2
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    • pp.77-82
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    • 2006
  • An efficient transformation system for the production of transgenic plants has been developed for tall fescue (Festuca arundinacea Schreb.) via Agrobacterium-mediated transformation of seed-derived callus. From the point of morphogenetic capacity, three types of callus were selected. High frequency of plant regeneration was obtained by selection of type II callus, and the plant regeneration frequency was 52.6% when embryogenic callus were cultured on the regeneration medium. Supplementation of the media with 10 mg/L $AgNO_3$ and 40 mg/L cysteine enhanced frequencies of plant regeneration up to 65.3%. The highest transformation efficiency was also obtained when type II callus were inoculated with Agrobacterium. Southern blot analysis of PCR products of transgenic plants demonstrated that transgenes were successfully integrated into the genome of tall fescue. Efficient regeneration system and transformation established in this study will be useful for molecular breeding of tall fescue through genetic transformation.

Callus Induction and Plant Regeneration from Mature Seeds of Timothy (티모시 성숙종자로부터 캘러스 유도 및 식물체 재분화)

  • Lee, Ki-Won;Kim, Ki-Yong;Choi, Gi-Jun;Lim, Young-Chul;Kim, Won-Ho;Jung, Min-Wong;Seo, Sung;Lee, Byung-Hyun;Lee, Sang-Hoon
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.28 no.3
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    • pp.165-170
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    • 2008
  • Timothy (Phleum pratense L.) is an important grass species as forage. In order to optimize tissue culture conditions of timothy, the effects of plant growth regulators on callus induction and plant regeneration was investigated with mature seeds of colt cultivar. The optimal concentration of 2,4-D for the induction of primary callus from mature seeds was 3 mg/L. The highest embryogenic callus frequenc (25%) was observed when the mature seed were cultured on MS medium supplemented with 3 mg/L 2,4-D and 0.1 mg/L BA. The highest plant regeneration frequency was observed when type B callus was transferred to N6 medium supplemented with 1 mg/L 2,4-D and 3 mg/L BA. Regenerated plants were grown normally when shoots were transplanted to the soil. A short tissue culture period and regeneration system would be beneficial for molecular breeding of timothy by the production of transgenic plant.

Association of Novel Polymorphisms in Lymphoid Enhancer Binding Factor 1 (LEF-1) Gene with Number of Teats in Different Breeds of Pig

  • Xu, Ru-Xiang;Wei, Ning;Wang, Yu;Wang, Guo-Qiang;Yang, Gong-She;Pang, Wei-Jun
    • Asian-Australasian Journal of Animal Sciences
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    • v.27 no.9
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    • pp.1254-1262
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    • 2014
  • Lymphoid enhancer binding factor 1 (LEF-1) is a member of the T-cell specific factor (TCF) family, which plays a key role in the development of breast endothelial cells. Moreover, LEF-1 gene has been identified as a candidate gene for teat number trait. In the present study, we detected two novel mutations (NC_010450.3:g. 99514A>G, 119846C>T) by DNA sequencing and polymerase chain reaction-restriction fragment length polymorphism in exon 4 and intron 9 of LEF-1 in Guanzhong Black, Hanjiang Black, Bamei and Large White pigs. Furthermore, we analyzed the association between the genetic variations with teat number trait in these breeds. The 99514A>G mutation showed an extremely significant statistical relevance between different genotypes and teat number trait in Guanzhong (p<0.001) and Large White (p = 0.002), and significant relevance in Hanjiang (p = 0.017); the 119846C>T mutation suggested significant association in Guanzhong Black pigs (p = 0.042) and Large White pigs (p = 0.003). The individuals with "AG" or "GG" genotype displayed more teat numbers than those with "AA"; the individuals with "TC" or "CC" genotype showed more teat numbers than those with "TT". Our findings suggested that the 99514A>G and 119846C>T mutations of LEF-1 affected porcine teat number trait and could be used in breeding strategies to accelerate porcine teat number trait improvement of indigenous pigs breeds through molecular marker assisted selection.