발아종자의 분열조직을 이용한 효율적인 콩 형질전환 방법
Efficient Transformation Method of Soybean Using Meristematic Tissues of Germinating Seeds
2
조회
0
다운로드
4
인용
0
참고문헌
키워드
초록
An efficient transformation method for soybean [Glycine max (L.) Merr.] using meristematic tissues of germinating seeds has been established. The embryonic axes were excised from germinating seeds of Korean soybean cultivar, Iksannamulkong and 0.5-2 cm long segment containing meristematic tissues were prepared by cutting hypocotyl region. The explants were inoculated with Agrobacterium tumefaciens strain LBA4404 harboring a binary vector with the bar gene as a selectable marker gene and a β-glucuronidase (GUSINT) reporter gene, and then co-cultured for 7 days on co-cultivation medium (CCM). The meristematic tissues were cultured on shoot induction medium (SIMP6) supplemented with 0.4 mg/l N6-benzylaminopurine (BAP) and 0.1 mg/l indolebutyric acid (IBA) in the presence of 6 mg/l L-phosphinotricin (PPT) for 2 weeks and the surviving explants were transferred to shoot elongation medium (SEMP6). Transformation was confirmed by Southern blot analysis and the transformation efficiencies ranged from 1.48 to 2.07%. The new modified transformation method was successfully implemented for obtaining several transgenic lines with SMV-CP gene. It is expected that this method could efficiently be used for the transformation of recalcitrant soybean cultivars.
Abstract
An efficient transformation method for soybean [Glycine max (L.) Merr.] using meristematic tissues of germinating seeds has been established. The embryonic axes were excised from germinating seeds of Korean soybean cultivar, Iksannamulkong and 0.5-2 cm long segment containing meristematic tissues were prepared by cutting hypocotyl region. The explants were inoculated with Agrobacterium tumefaciens strain LBA4404 harboring a binary vector with the bar gene as a selectable marker gene and a β-glucuronidase (GUSINT) reporter gene, and then co-cultured for 7 days on co-cultivation medium (CCM). The meristematic tissues were cultured on shoot induction medium (SIMP6) supplemented with 0.4 mg/l N6-benzylaminopurine (BAP) and 0.1 mg/l indolebutyric acid (IBA) in the presence of 6 mg/l L-phosphinotricin (PPT) for 2 weeks and the surviving explants were transferred to shoot elongation medium (SEMP6). Transformation was confirmed by Southern blot analysis and the transformation efficiencies ranged from 1.48 to 2.07%. The new modified transformation method was successfully implemented for obtaining several transgenic lines with SMV-CP gene. It is expected that this method could efficiently be used for the transformation of recalcitrant soybean cultivars.
참고문헌 (26)
- Buhr T, "Nuclear localization of RNA transcripts downregulate seed fatty acid genes in transgenic soybean", Plant J, 30, pp.155~163 (2002)
- Cheng T-Y, "Plant regeneration from soybean. cotyledonary node segments in culture", Plant Sci. Lett, 19, pp.91~99 (1980)
- De block M, "The cell biology of plant transformation: Current state, problems, prospects and the implications for the plant breeding", Euphytica, 71, pp.1~14 (1993)
- Finer JJ, "Transformation of soybean via particle bombardment of embryogenic suspension culture tissue", In Vitro Cell and Develop Biol-Plant, 27P, pp.175~182 (1991)
- Franklin G, "Factors influencing regeneration of soybean from mature and immature cotyledons", Plant Growth Regul, 43, pp.73~79 (2004)
- Gamborg OL, "Nutrient requirement of suspensions cultures of soybean root cells", Exp. Cell Res, 50, pp.151~157 (1968)
- Hari B, Krishna, "Engineering soybean for enhanced sulfur amino acid content", Crop Sci, 45, pp.454~461 (2005)
- Hinchee MAW, "Production of transgenic soybean plants using Agrobacterium-mediated DNA transfer", Bio, 1988, pp.6, 915~922
- Kimball SL, "Adventitious bud development of soybean. hypocotyl sections in culture", Crop Sci, 13, pp.758~760 (1973)
- 권순종, "아그로박테리움을 이용한 제초제 저항성 콩 개발", Journal of Plant Biotechnology, 35(1), pp.69~74 (2008), 한국식물생명공학회
- Liu, HK, "Efficient Agrobacterium tumefaciens-mediated transformation of soybeans using an embryonic tip regeneration system", Planta, 219, pp.1042~1049 (2004)
- Macrae TC, "Laboratory and field evaluations of transgenic soybean exhibiting high-dose expression of a synthetic Bacillus thuringiensis cry1A gene for control of Lepidoptera", J. Econ. Entomol, 98(2), pp.577~587 (2005)
- Mante, S, "A simple, rapid protocol for adventitious shoot development from mature cotyledons of Glycine max cv Bragg", In Vitro Cell. Dev. Biol, 25, pp.385~388 (1989)
- McCabe, DE, "Stable transformation of soybean (Glycine max) by particle acceleration", Bio, 1988, pp.6, 923~926
- Micheal D, "Current use of transgenic herbicide-resistant soybean and corn in the USA", Crop Protection, 19, pp.765~771 (2000)
- Murashige T, "A revised medium for rapid growth and bioassays with tobacco tissue cultures", Physiol. Plant, 15, pp.473~479 (1962)
- Olhoft PM, "The role of thiol compounds in increasing Agrobacterium-mediated transformation of soybean cotyledonarynode cells", Plant Cell Rep, 20, pp.731~737 (2001)
- Paz MM, "Improved cotyledonary node method using an alternative explant derived from mature seed for efficient Agrobacterium-mediated soybean transformation", Plant Cell Rep, 25, pp.206~213 (2006)
- Rogers S, "Extraction of total DNA from plants, algae and fungi In: Plant Molecular Biology Manual", pp.1~8 (1994), The Netherlands, Kluwer Academic Publisher
- Sairam RV, "A study on the effect of genotypes, plant growth regulators and sugars in promoting plant regeneration via organogenesis from soybean cotyledonary nodal callus", Plant Cell Tiss. Organ Cult, 75, pp.79~85 (2003)
- Saka H, "Stimulation of multiple shoot formation on soybean stem nodes in culture", Plant Sci. Lett, 19, pp.193~201 (1980)
- Shan Z, "Development of a highly efficient, repetitive system of organogenesis in soybean (Glycine max (L.) Merr)", Plant Cell Rep, 24, pp.507~512 (2005)
- Jang-Yong Lee, "An Efficient Selection Scheme for Agrobacterium-mediated Co-transformation of Rice Using Two Selectable Marker Genes hpt and bar", 한국육종학회지, 38(3), pp.173~179 (2006), 한국육종학회
- Trick HN, "SAAT:Sonication Assisted Agrobacterium-mediated Transformation", Transgenic Research, 6, pp.329~336 (1997)
- Wang G, "Hypocotyl-based Agrobacteriummediated transformation of soybean (Glycine max) and application for RNA interference", Plant Cell Rep, 27, pp.1177~1184 (2008)
- Wright, MS, "Initiation and propagation of Glycine max L. Merr.:Plants from tissue-cultured epicotyls", Plant Cell Tissue and Organ Cult, 8, pp.83~90 (1987)