Korean J. Plant Res. 27(1):043-050(2014) http://dx.doi.org/10.7732/kjpr.2014.27.1.043 Print ISSN 1226-3591 Online ISSN 2287-8203 Original Research Article 녹색꽃잎도라지 의기관분화에미치는배지조성및생장조절제의영향 권수정 1, 조갑연 2, 김학현 1 * 1 우송정보대학플라워코디 조경과, 2 우송정보대학식품영양조리계열 Medium Composition and Growth Regulator on Organogenesis Platycodon grandiflorum (Jacq.) A. DC. with Yellow Green Petals Soo Jeong Kwon 1, Kab Yeon Cho 2 and Hag Hyun Kim 1 * 1 Department of Flower Floral Plant Coordi & Landscape Architecture, Woosong College, Daejeon 300-715, Korea 2 Department of Food Nutrition and Cookery, Woosong College, Daejeon 300-715, Korea Abstract - This study was carried out to determine the optimal medium composition and growth regulators for the micropropagation of Platycodon grandiflorum (Jacq.) A. DC. Nodes containing yellow green petals were used as plant materials to execute the study. The best performance of adventitious root development was found in 1/4 strength of MS basal salt and the growth was satisfactory in the concentration of 1/2 MS medium. The best condition for adventitious root development and growth was observed in the higher concentration (5%) of sucrose and activated charcoal free 1/4MS medium respectively. Adventitious roots were developed at the controlled culture medium at ph 4.8 with a tendency of suppression with higher levels of ph. However, it was prevailed that the development and growth depended on the concentration of agar. The lower concentration of agar (0.4%) was performed better than that of higher concentration (1.2%), whereas the agar concentration (0.4%) showed the best performance for the development and growth of adventitious roots. For the development of shoots containing node, BA combined with IAA was more effective than kinetin with IAA or NAA. The highest shoot development (3.9 shoots per explant) was performed on MS medium supplemented with 0.1 mg /L BA and 0.5 mg /L IAA. Key words - Organogenesis, Adventitious root, Medium Composition, Culture Materials, Micropropagation, Sucrose 서언 도라지는초롱꽃과, 초롱꽃속에속하는다년생숙근초로서세계의온대및아열대지역에약 60속 1,500 종이분포되어있다. 그중우리나라에 9속 50여종이서식하고있고, 그대표적인것으로는백도라지, 청도라지, 아기도라지, 좀도라지등이있다 (Ahn et al., 1986; Cho, 1984). 우리나라에서는산야에자생하는것을채취이용하여왔으나최근에는수요가증가하여, 일반농가에서재배가늘어가고있다. 도라지는단백질, 지질, 당류, 전분, 철분, saponin, inulin, phytosterin, platycodinin 등을함유하고있는식물로서흰색 * 교신저자 (E-mail) : hkyushu@hanmail.net 의뿌리는다육질로식용 약용으로많은양이재배되고있다. 꽃색은백색, 자색이나원예종으로분홍색이있으며꽃모양은홑꽃및겹꽃으로아름답게개화되어정원, 조경용, 절화용화훼로이용되어지상부와지하부를모두이용하는유망한자생식물이다. 본연구실에서자체육성한돌연변이종인 녹색꽃잎도라지 는화색이엷은녹색으로, 일반종에비해개화기간이길고화색의희소성에의해관상가치가높은장점이있는반면불임성으로종자가형성되지않아번식에어려움이있다. 조직배양은여러조직으로부터균일한식물체를단기간내에대량생산할수있는하나의방법으로이용되어져왔다. 이러한조직배양은사용되는식물체의절편부위에따라다세포기원의부정아가형성되는경우와단세포기원의체세포배가배 c 본학회지의저작권은 ( 사 ) 한국자원식물학회지에있으며, 이의무단전재나복제를금합니다. This is an Open-Access article c 본 distributed 학회지의 under 저작권은 the terms ( of 사 the ) 한국자원식물학회지에 Creative Commons -43- Attribution 있으며 Non-Commercial, 이의무단전재나 License (http://creativecommons.org/licenses/by-nc/3.0) 복제를금합니다. which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
韓資植誌 Korean J. Plant Res. 27(1) : 043~050(2014) 발생캘러스를통하거나조직편에서직접발생하여식물체로재분화됨으로서농작물개량및급속증식에효율적인방법으로이용될수있다 (Hussey and Stacey, 1981). 또한, 조직배양기술이여러분야에응용되기위해서는캘러스유기및세포배양등을통한중요한역할을하게된이후기내에서식물체를재분화시키고자여러가지방법이이용되고있다 (Amirouche et al., 1985; Hussey and Staecy, 1981; Kim et al., 2003). 본실험은관상가치가높아원예품종화가가능할것으로판단되는 녹색꽃잎도라지 의대량증식을위한기초자료를얻을목적으로, 기내배양시, 식물체의재분화에미치는배지구성물질및생장조절제의영향에대하여조사하였다. 재료및방법식물재료및처리내용시험재료는기내에서배양중인 녹색꽃잎도라지 (Fig. 1) 의마디를포함한줄기절편을 1 cm길이로잘라배지에치상하였다. 배양은 petri dish 상에서반복당 6절편으로하였으며완전임의배치법 3반복으로비교분석하였다. MS 배지 (Murashige and Skoog, 1962) 구성물질의적정농도실험 (2MS, MS, 1/2MS, 1/4MS 배지 ) 은 sucrose 3%, ph 5.8로조절한후 agar 0.8% 를첨가하였으며, sucrose 실험 (1, 3, 5, 7%) 은 MS배지를기본으로 ph 5.8 로조절한후 agar 0.8% 를첨가하였다. 활성탄농도실험은 MS배지와대조구 (1/4MS 에 sucrose 5% 를첨가한배지 ) 의경우활성탄을첨가하지않았으며, 이외처리구는각각 0.05, 0.1, 0.5, 1.0% 첨가하였다. ph (3.8, 4.8, 5.8, 6.8, 7.8) 및 agar 농도실험 (0.4, 0.6, 0.8, 1.0 1.2%) 은 MS배지구성물질농도, sucrose 농도및 ph 실험의결과를바탕으로 sucrose 5% 를첨가하고 ph 5.8 로조절한후 agar 0.8% 를첨가한 1/4MS 배지에 4주 간배양하였다. 또한, 생장조절제실험의경우, 기본배지는배양실험결과가장좋았던 1/4MS 배지를기본으로 sucrose 5% 를첨가한후, ph 4.8로조절한배지에 agar 0.4% 를첨가하였고예비실험결과가장좋았던 cytokinin 류인 BA 0.1 mg /L 와 kinetin 0.5 mg /L를 auxin 류인 NAA 와 IAA 0.1, 0.5, 1, 5, 10 mg /L 와혼용첨가하여 8주간배양하였다. 모든실험의배양조건은 25 ± 1, 40 μmol ㆍm -2 ㆍs -1 의광으로 16시간조명하였으며, 신초및부정근의수와길이등을조사하였다. 결과및고찰배지무기염류의농도효과 녹색꽃잎도라지 의기관분화에적합한배지구성물질의농도를알아보기위하여 2MS, MS, 1/2MS 및 1/4MS 배지에배양해본결과는 Fig. 2와같다. 신초의형성은처리구간에큰차이를보이지않았으며, 생장은 2MS 배지에서가장왕성하였다. 부정근의형성은배지의무기물농도가낮을수록양호한형성을보여, 1/4MS 배지에서가장좋았다. 그러나무기물의농도가가장높았던 2MS 배지에서는부정근의형성이관찰되지않았다. 부정근의생장도무기물의농도가낮은 1/2MS 와 1/4MS 배지에서양호한결과를나타냈다. 배지의무기물농도는기관분화및식물체생장에영향을미치는데, 고추냉이의경우신초증식은무기성분의농도에비례하며 (Park et al., 2007), 감자는저농도의무기성분이신초의생장 (Evans, 1993) 에흰꿀풀은부정근형성 (Kwon et al., 2012) 에효과적인것으로알려져있다. 본실험의결과녹색꽃잎도라지의신초의생장과부정근의형성및생장은무기물의농도에따라그효과가다른것으로나타나, 각기관분화및생장에따른 MS 무기물농도에대한요구도는차이가있는것으로생각되었다. Fig. 1. Plant material used in the study. Sucrose 농도효과배지에공급되는탄소원으로가장많이사용되고있는 sucrose 가기관분화에미치는효과를알아보기위해 1, 3, 5, 7% 의농도로첨가하여배양한결과는 Fig. 3과같다. 신초의형성은처리구간에큰차이를보이지않았으나, 신초의생장은고농도의 sucrose를첨가한구에서양호한경향으로, 특히 5% 첨가구에서절편체당 1.6 cm로가장좋았다. 부정근또한고농도구에서양호한경향을보여 sucrose 5% 첨가에서절편체당 8.7개로가장많은부정근이형성되었으며, 생장도왕성하였다. 그러나본 -44-
녹색꽃잎도라지 의기관분화에미치는배지조성및생장조절제의영향 Fig. 2. Effect of culture media on shoot and adventitious root formation from node of Platycodon grandiflorum (Jacq.) A. DC. with yellow green petal for 4 weeks in culture. Each bar represents the mean ± SE of triplicate experiments. Fig. 3. Effect of sucrose concentration on shoot and adventitious root formation from node of Platycodon grandiflorum (Jacq.) A. DC. with yellow green petal for 4 weeks in culture. Each bar represents the mean ± SE of triplicate experiments. -45-
韓資植誌 Korean J. Plant Res. 27(1) : 043~050(2014) 실험에서배지내당농도가가장높았던 7% 첨가구는생장이저하되는것으로나타났다. 이는 홍경천 의경우 3% 농도에서높은부정근의유도율을보인다 (Bae et al., 2009) 는보고와차이를보인반면, 5% 농도의 sucrose 첨가에의해고구마의부정근수가증가하였다 (Jarret and Gawel, 1991) 는결과와유사하였다. 이같은결과는조직배양시배양체의발근에필수적인요소이며 (Gautheret, 1969), 식물의형태형성에가장중요한요소 (Romano et al., 1995) 로알려진배지내의 sucrose 는농도의고저및품종에따라기관분화에대한반응이상이한것으로판단되었다. 녹색꽃잎도라지 의경우, 신초의생장및부정근의형성을위해기본 MS배지에첨가한 sucrose 의농도에비해조금높은농도의첨가가필요할것으로생각되었다. 활성탄 (activated charcoal) 농도효과 녹색꽃잎도라지 의기관분화에미치는활성탄의효과를알아보기위해 MS, Cont. (1/4MS, sucrose 5%), 0.05, 0.1, 0.5, 1.0% 의농도로첨가하여배양한결과는 Fig. 4와같다. 절을배양재료로활성탄첨가의유무에따른신초의형성은, 처리구간에큰차이를보이지않았던반면, 부정근의형성은무첨가구에비해첨가구에서절편체당 1.3 개이하의낮은형성을나타내, 활성탄의첨가는부정근의형성을억제시키는요인으로생각되었다. 특히고농도구인 0.5, 1.0% 첨가구에서는 0.4, 0.1 개의미미한부정근의형성을보였다. 생장또한활성탄의무첨가구에서양호한결과를나타냈다. 일반적으로활성탄은배양절편으로부터유출되는독성물질을흡수하고배양액의갈변화를방지하여생존율을높이며배형성이나뿌리의발생을촉진시킨다고알려져있다 (Teng, 1997). 그러나본실험의경우, 활성탄은뿌리의발생에오히려억제적으로작용한것으로보여품종에따라뿌리형성에미치는활성탄의효과가다르게나타난다는것을알수있었다. 또한, 활성탄을첨가하지않은 MS배지에서는다른처리구들과달리신초의생장은양호하였던반면, 부정근은전혀형성되지않았다. 이상의결과로부터 녹색꽃잎도라 Fig. 4. Effect of activated charcoal concentration on shoot and adventitious root formation from node of Platycodon grandiflorum (Jacq.) A. DC. with yellow green petal for 4 weeks in culture. Each bar represents the mean ± SE of triplicate experiments. MS : MS basal medium (Sucrose 3%, agar 0.8%, ph 5.8) / Cont. : 1/4MS medium (Sucrose 5%, agar 0.8%, ph 5.8). -46-
녹색꽃잎도라지 의기관분화에미치는배지조성및생장조절제의영향 지 의부정근형성은활성탄의첨가유무뿐만아니라, 배지의무기물농도에큰영향을받는다는것을알수있었다. 지의적정 ph는각식물체간에많은차이가있으며, 녹색꽃잎도라지 의적정 ph는 4.8로판단되었다. ph 효과 녹색꽃잎도라지 의기관분화에미치는 ph의효과를알아보기위하여 ph 범위를 3.8~7.8로조절한배지에배양한결과는 Fig. 5와같다. 신초의형성은모든처리구에서큰차이를보이지않았던반면, 신초의생장은 ph 4.8 로조절한배지에서가장좋았으며, 배지의 ph가높아질수록저조한생장을나타냈다. 부정근의형성및생장또한신초의생장과유사한결과로 ph 4.8 의배지에서부정근의수와길이가절편체당 4.3개, 1.2 cm로가장양호하였으나, ph 가높아질수록생장은저조한경향을보였다. 식물조직배양에사용되는배지의일반적인 ph 범위는주로 5.5~5.8이며, 도라지의조직배양에있어서도대부분 ph 5.3~5.8의범위에서이루어지고있어 (Ko et al., 1993; Chung et al., 2002; Choi et al., 2005), 본실험의결과에서보여준적정 ph 범위와조금차이가있었다. 그러나식물재료에따라배양에적합한 ph 범위는다양하여 Anderson (1975) 은 rhododendron을이용한신초증식에서적합한 ph는 4.5라고하였으며, Yang et al. (1997) 은독말풀모상근의성장률이 ph 6.3에서증가되었다고보고하였다. 이같은결과들을볼때배 Agar 의농도효과 녹색꽃잎도라지 의기관분화에미치는 agar 농도의효과를알아보기위하여 0.4, 0.6, 0.8, 1.0, 1.2% 의농도로첨가하여배양한결과는 Fig. 6과같다. 신초의형성은 agar의농도가 0.6% 인배지에서절편체당 2.4개, 생장은저농도구인 0.4% 배지에서 1.1 cm로가장좋은결과를보였는데저농도의 agar 를첨가한경우, 절편체로부터분비되는독성물질이배지로확산이잘되며, 배지내의영양분의구배가적어생육을촉진한결과로생각되었다. 부정근의경우는배지의견고도가낮을수록양호한결과를나타냈는데특히, 부정근의형성과생장은 0.4% 의농도구에서각각 4.2개, 1.0 cm로가장높은결과를보였던것에비해, agar의농도가가장높은 1.2% 첨가구에서절편체당 0.3 개로부정근의형성은억제되는것으로나타났으며, 생장또한 0.1 cm로저조하였다. 생장조절제효과 녹색꽃잎도라지 의기관분화에미치는생장조절제의종류및농도를알아보고자 BA (0.1 mg /L), kinetin (0.5 mg /L), IAA Fig. 5. Effect of ph on shoot and adventitious root formation from node of Platycodon grandiflorum (Jacq.) A. DC. with yellow green petal for 4 weeks in culture. Each bar represents the mean ± SE of triplicate experiments. -47-
韓資植誌 Korean J. Plant Res. 27(1) : 043~050(2014) Fig. 6. Effect of agar concentration on shoot and adventitious root formation from node of Platycodon grandiflorum (Jacq.) A. DC. with yellow green petal for 4 weeks in culture. Each bar represents the mean ± SE of triplicate experiments. 및 NAA (0.1, 0.5, 1, 5, 10 mg /L) 를각각혼용처리하여배양한결과는 Table 1, 2와같다. 마디를포함한줄기절편을배양재료로하여배양한경우마디부위에서직접신초가형성되는것이관찰되었는데 BA와 NAA 의혼용처리구에서의신초의형성및생장은 NAA 의농도가높을수록감소하는경향을보였던반면대조구 ( 무첨가구 ) 및 NAA 의농도가낮을수록양호한결과를보였다. 부정근의형성및생장또한처리농도에관계없이대조구에비해전반적으로저조한결과를보였으며, 특히 NAA 의농도가높을수록억제되는것으로나타나, NAA 의첨가는 녹색꽃잎도라지 의부정근형성및생장을저해하는생장조절물질로생각되었다. 캘러스는 NAA 1 mg /L 농도이상의처리구에서많이유기되었으며, BA 0.1 mg /L와 NAA 5 mg /L 혼용첨가한배지에서가장많은캘러스유기를보였다. BA와 IAA 의혼용첨가에따른신초의형성은 BA 0.1 mg /L와 IAA 0.5 mg /L 혼용구에서양호한결과를나타내었던반면, 신초의생장은 BA와 IAA 를첨가한배지보다무첨가한구에서가장왕성하였다. 부정근또한 NAA 보다는 IAA 의첨가에의해형성이촉진되었으며 BA 0.1 mg /L와 IAA 0.5 mg /L혼용구에서절편체당 14.3 개로가장많은부정근이형성되었다. 부정근의생장은무첨가보다 IAA 첨가구에서대체적으로왕성하였다 (Table 1). Kothari and Chandra (1984) 은 Africa marigold 의배양에서 shoot 의형성이 BA와 IAA 를첨가한배지에서많은분화를 나타내었고 Billings et al. (1988) 은 blueberry 의엽절편배양에서 NAA 를첨가한경우캘러스형성이촉진되고 shoot 의분화가억제되었다고보고하였다. 또한 Moore (1986) 는감귤류대목의절간절편배양에서 shoot 의유도는 BA의단용이나 NAA 와의혼용첨가가좋으나장기간배양에서 NAA 는 shoot형성을억제하였다는보고와유사한결과로 NAA 는녹색꽃잎도라지의 shoot 형성에적합하지않은생장조절제로생각되었다. Kinetin 과 NAA 의혼용처리구에있어신초의형성은처리구간에큰차이는없었으나 kinetin 0.5 mg /L와 NAA 10 mg /L혼용첨가구에조금저조한형성을보였으며, 신초의생장은 NAA 무첨가구에서가장왕성한생육을나타냈다 (Table 2). 부정근의형성및생장은 kinetin 0.5 mg /L와 NAA 0.1 mg /L 혼용첨가구에서가장높은결과를나타내었으나, NAA 의농도가 5 mg /L 이상첨가한경우, 부정근의생장은저조한경향을보였다. Kinetin 과 IAA 의혼용은 IAA 0.1 과 0.5 mg /L를혼용첨가한구에서가장양호한신초의형성을보였으며, 나머지처리구는대조구와처리구간차이가없었으며, 신초의생장은 IAA 농도에관계없이무첨가배지에비해낮은결과를나타냈다. 부정근의형성은 kinetin 과 IAA 를첨가함에따라감소하였으며농도가높아질수록저조한형성을나타낸반면생장은 kinetin 0.5 mg /L와 IAA 5 mg /L를혼용한구에서절편체당 5.9 cm로가장왕성한부정근의생장을보였다. -48-
녹색꽃잎도라지 의기관분화에미치는배지조성및생장조절제의영향 Table 1. Effect of BA, NAA and IAA on organogenesis from node of Platycodon grandiflorum (Jacq.) A. DC. with yellow green petal cultured on 1/4MS medium for 8 weeks BA NAA IAA ( mg /L) No. of shoots Shoot length ( cm ) No. of roots Root length ( cm ) Callus wt. ( mg ) Control 2.1 ± 0.1 z 2.6 ± 0.2 8.4 ± 0.7 4.7 ± 0.4 5.0 ± 1.3 0.1 2.3 ± 0.2 1.4 ± 0.1 4.3 ± 0.9 4.3 ± 0.9 74.7 ± 17.5 0.5 1.8 ± 0.1 0.9 ± 0.1 5.1 ± 1.4 3.3 ± 0.8 66.8 ± 8.9 0.1 1 1.5 ± 0.2 0.6 ± 0.1 6.9 ± 1.2 4.1 ± 0.7 200.5 ± 43.6 5 1.6 ± 0.2 0.5 ± 0.1 5.1 ± 0.9 0.7 ± 0.1 219.5 ± 27.1 10 0.7 ± 0.2 0.2 ± 0.1 5.9 ± 1.1 0.6 ± 0.1 181.7 ± 26.3 0.1 2.5 ± 0.3 1.8 ± 0.1 7.1 ± 0.8 6.3 ± 0.6 24.6 ± 10.9 0.5 3.9 ± 0.5 1.4 ± 0.1 15.1 ± 2.3 6.6 ± 0.5 45.9 ± 13.4 0.1 1 2.2 ± 0.2 1.4 ± 0.1 8.1 ± 1.1 6.6 ± 0.6 66.5 ± 16.2 5 2.1 ± 0.2 2.0 ± 0.2 8.1 ± 1.2 6.7 ± 0.8 38.6 ± 15.7 10 1.9 ± 0.2 2.0 ± 0.2 8.0 ± 1.0 4.9 ± 0.6 52.7 ± 14.3 z mean of 20 plants ± S.E. Table 2. Effect of kinetin, NAA and IAA on organogenesis from node of Platycodon grandiflorum (Jacq.) A. DC. with yellow green petal cultured on 1/4MS medium for 8 weeks Kinetin NAA IAA ( mg /L) No. of shoots Shoot length ( cm ) No. of roots Root length ( cm ) Callus wt. ( mg ) Control 2.1 ± 0.1 z 2.6 ± 0.2 7.9 ± 0.7 4.7 ± 0.4 5.0 ± 1.3 0.1 2.2 ± 0.2 2.3 ± 0.3 8.2 ± 0.2 5.6 ± 0.8 50.3 ± 12.3 0.5 2.1 ± 0.1 1.6 ± 0.1 8.0 ± 0.2 5.4 ± 0.4 114.2 ± 14.9 0.5 1 2.1 ± 0.1 0.9 ± 0.1 9.1 ± 1.1 5.1 ± 0.5 114.5 ± 11.8 5 2.2 ± 0.2 0.7 ± 0.1 6.8 ± 1.7 1.6 ± 0.4 205.8 ± 15.3 10 1.2 ± 0.2 0.4 ± 0.1 6.9 ± 1.3 0.9 ± 0.1 223.9 ± 29.9 0.1 2.6 ± 0.3 1.9 ± 0.1 7.5 ± 1.5 5.0 ± 0.7 52.0 ± 13.5 0.5 2.7 ± 0.4 1.7 ± 0.1 6.4 ± 0.9 5.9 ± 0.7 45.1 ± 9.5 0.5 1 2.1 ± 0.1 2.2 ± 0.2 5.9 ± 1.0 4.6 ± 0.6 37.9 ± 10.9 5 1.9 ± 0.1 2.2 ± 0.2 5.9 ± 0.8 4.6 ± 0.6 37.8 ± 9.1 10 2.1 ± 0.1 2.0 ± 0.2 6.0 ± 0.8 4.7 ± 0.8 73.8 ± 18.4 z mean of 20 plants ± S.E. 캘러스의유기는모든혼용첨가구에서관찰되었으며, 특히 NAA 를고농도로첨가한구에서가장많은캘러스가유기되었다. 적요본연구는 녹색꽃잎도라지 의기내배양시배지구성물질의적정농도구명에의한대량번식을목적으로실시하였다. 녹색꽃잎도라지 의절을배양재료로배양조건은 MS배지의여러가지구성물질의농도를달리한결과, 1/4MS 배지에서가장양 호한부정근의형성을보였으나생장은 1/2MS 배지에서좋았다. Sucrose 첨가는농도가높을수록신초와부정근의형성및생장이좋았다. 활성탄은무첨가구에서가장많은부정근의형성과양호한생장을보였다. 배지의 ph는 4.8로조절된배지에서가장많은부정근을형성하였으며, ph 가높아질수록그형성은낮아지는경향을보였고, 부정근과신초의생장또한 ph 4.8 에서가장왕성하였다. Agar 농도별실험에서부정근의형성과생장은그농도가낮아질수록양호한경향을보여가장낮은첨가구인 0.4% 농도구에서가장많은부정근의형성과왕성한생 -49-
韓資植誌 Korean J. Plant Res. 27(1) : 043~050(2014) 장을보였다. 생장조절제를혼용첨가한경우신초의형성은 BA 와 IAA 의혼용구가 kinetin 과 IAA 또는 NAA 혼용구에비해효과적이었으며, BA 0.1 mg /L와 IAA 0.5 mg /L혼용구에서절편체당 3.9개로가장많은신초가형성되었다. 사사 본연구는농림축산식품부고부가가치식품기술개발사업 (112076-03-2-HD020) 에의해이루어진것임. References Ahn, C.S., S.J. Chung, G.C. Koh and S.H. Park. 1986. Isolation and culture of protoplasts from Platycodon grandiflorum (Jacq.) A. DC. and Codonopsis lanceolata (S. et Z.) Trautv. J. Kor. Soc. Hort. Sci. 27(3):205-212. Amirouche, L., T. Stuchbury and S. Matthews. 1985. Comparisons of cultivar performance on different nutrient media in a routine method for potato micropropagation. Potato Res. 28:469-478. Anderson, W.C. 1975. Production of rhododendrons by tissue culture. Ⅰ. Development of a culture medium for multiplication of shoots. Proc. Int. Plant Prop. Soc. 25:129-135. Bae, K.H., E.S. Yoon and Y.E. Choi. 2009. In vitro culture of adventitious root from Rhodiola sachalinensis. Korean J. Plant Res. 22(4):281-286 (in Korean). Billings, S.G., C.K. Chin and G. Jelenkovic. 1988. Regeneration of blueberry plantlets from leaf segments. HortScience 23:763-766. Cho, J.T. 1984. Physiological and ecological studies on the Chinese bellflower, Platycodon grandiflorum (Jacq.) A. DC. Ⅰ. Studies on seed germination, growth and flowering of Chinese bellflower. J. Kor. Soc. Hort. Sci. 25(3):187-193. Choi, S.R., M.J. Kim, J.S. Eun, M.S. Ahn, H.C. Lim and J. Ryu. 2005. The growth response of balloon flower (Platycodon grandiflorum A. DC.) plantlets in vitro as affected by air exchanges and light intensity. Korean J. Plant Biotechnol. 32(1):23-29 (in Korean). Chung, J.H. and S.H. Cho. 2002. Tissue cultures of Platycodon grandiflours DC. J. Agriculture & Life Sciences 36(4):9-18 (in Korean). Evans, N.E. 1993. A preliminary study on the effects of nitrogen supply on the growth in vitro of 9 potato genotypes (Solanum spp.) J. Expt. Bot. 44:837-841. Gautheret, R.J. 1969. Investigations on the root formation in the tissue of Heiianthus tuberosus cultured in vitro. Amer. J. Bot. 56:702-717. Hussey, G. and N.J. Stacey. 1981. In vitro propagation of potato (Solanum tuberosum L.). Ann. Bot. 48:787-796. Jarret, R.L. and N. Gawel. 1991. Chemical and environmental growth regulation of sweet potato (Ipomoea batatas (L.) Lam.) in vitro. Plant Cell Tiss. Organ Cult. 25:153-159. Kim, I.H., M.A. Kim, H.H. Kim and C.H. Lee. 2000. Effect of culture materials and growth regulator on plant regeneration of Platycodon grandiflorum and Codonopsis lanceolata in vitro. Korean J. Plant Res. 92-93 (Abstr.) (in Korean). Ko, J.A., Y.S. Kim, M.J. Kim and J.S. Eun. 1993. Somatic embryogenesis and plant regeneration from leaf and stem culture of Platycodon grandiflorum. Korean J. Breed. 25(4):337-343 (in Korean). Kothari, S.L. and N. Chandra. 1984. In vitro propagation of African marigold. HortScience 19:703-705. Kwon, S.J., J.H. Park, S.H. Woo and H.H. Kim. 2012. Effect of media components on organogenesis of tetraploid in Prunella vulgaris for. albiflora. Korean J. Plant Res. 42 (Abstr.) (in Korean). Moore, G.A. 1986. In vitro porpagation of citrus root stock. Hortscience 21:300-301. Murashige T. and F. Skoog. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15:473-497. Park, Y.Y., M.S. Cho and J.B. Chung. 2007. Effect of salt strength, sucrose concentration and NH 4/NO 3 ratio of medium on the shoot growth of Wasabia japonica in vitro culture. J. Plant Biotechnol. 34(3):263-269 (in Korean). Romano, A., C. Noronha and M.A. Martins-Loucao. 1995. Role of carbohydartes in micropropagation of cork oak. Plant Cell Tiss. Organ Cult. 40:159-167. Teng, W.L. 1997. Activated charcoal affects morphogenesis and enhances sporophyte regeneration during leaf cell suspension during leaf cell suspension culture of Platycerium bifurcatum. Plant Cell Rep. 17:77-83. Yang, D.C., H.M. Kang, K.S. Lee, Y.H. Kim and D.C. Yang. 1997. Effects of ph, sucrose and vitamins on the growth and tropane alkaloid production of hairy roots of Datura stramonium var. tatula Torr. Korean J. Plant Tissue Culture 24(3):143-148 (in Korean). (Received 11 June 2013 ; Revised 4 October 2013 ; Accepted 27 November 2013) -50-