Water lettuce (Pistia stratiotes L.) is a free-floating perennial herbaceous plant with rosette leaves and a stem. Although this plant multiplies and has adverse effects on aquatic ecosystems, it can be used for biological purification of polluted water and production of valuable substances as a traditional medicine. In this study, we report a protocol to establish an in vitro micropropagation method based on direct shoot organogenesis from stem explants. In media comprising two types of basal medium and different growth regulators, multiple shoot organogenesis was observed on stems. The micropropagation method was most effective on Schenk and Hildebrandt (SH) medium with 1 mg/L indole-3-acetic acid and 2 mg/L 6-benzylaminopurine supplemented with 30 g/L sucrose, on which all explants produced multiple shoots. The shoots rooted spontaneously on solid half-strength Murashige and Skoog (MS) and SH media without growth regulators (1/2MSO and 1/2SHO). However, roots developed more vigorously in liquid media. Regenerated plants colonized and grew more rapidly in SH basal medium than in MS basal medium and produced 6–8 stolons within 2 weeks on 1/2SHO. In summary, we established a method for micropropagation in vitro through direct organogenesis of water lettuce, which shows the potential of water lettuce as a model aquatic plant for phytochemical and pharmacological research.

This study was conducted to examine the effects of medium composition on organogenesis towards in-vitro cultured diploid and tetraploid Codonopsis lanceolata and obtain in-vitro mass propagation of superior species of C. lanceolata. Regarding MS medium composition for each concentration, diploid C. lanceolata was found to be declined. However, shoot and adventitious root formation were suppressed with higher mineral salt concentration, and active growth of shootand adventitious root was exhibited as 4.9 cm and 3.2 cm respectively in 1/2 MS medium. While in tetraploid C. lanceolata, it showed 2.9 cm and 3.2 cm respectively in 1/4 MS medium. In the case of sucrose concentration, no consistent decrease was observed for growth of shoot and adventitious root of diploid both at high and low concentration. The growth of shoot (at 3% concentration) and adventitious root (at 7% concentration) was 2.3 cm and 2.0 cm respectively. Although there was no difference in shoot formation of tetraploid C. lanceolata in all concentrations with the range of 1.7 ~ 1.8, there was a slight decrease in shoot growth at high concentration. Results revealed that the adventitious root formation was suppressed at high concentration. Concentration of agar exhibited no significant difference in shoot formation of diploid C. lanceolata at all concentrations. The highest result of adventitious growth (4.1 cm) was observed at 0.8% concentration. Slight inhibition of shoot formation and root formation of tetraploid C. lanceolata was observed at higher concentration. Shoot formation of diploid C. lanceolata also exhibited inhibition at higher concentration. Shoot formation of diploid C. lanceolata was increased at lower pH and shoot growth was the highest (2.3 cm) at pH 3.8. Adventitious root formation was higher at lower pH. Although there was no difference in shoot formation of tetraploid C. lanceolata presenting 1.7 ~ 1.8 regardless of high and low pH, growth inhibition was showed at higher pH. Adventitious root formation and growth showed a little higher result at pH 5.8.

미치광이풀 부정근으로부터 탈분화 및 재분화에 미치는 세포소기관의 형태 및 tropane alkaloids의 함량 변화를 TEM과 HPLC를 사용하여 조사하였다. Scopolamine and hyoscyamine 및 세포 소기관의 형태변화는 세포분화 및 탈분화 단계마다 차이를 보였다. 부정근은 배양 20일후에는 캘러스가 형성되었고, 배양 60일부터 줄기원기가 발생하였으며, 이후 완전한 줄기로 발달하였다. 부정근 정단부 조직을 TEM 분석 결과 핵, 액포, 전분체, 미토콘드리아, 골지체, 세포벽 등 다양한 세포소기관들이 형성되었으나, 배양 20일후부터는 소기관들이 적게 관찰되었다. 그러나 분화가 시작되는 배양 40일 후 callus 세포에는 다수의 전분체 등 다양한 세포소기관들이 다시 관찰되었고, 배양 60일 이후에는 다시 전분립의 수가 감소하였다. 배양 80일째 세포는 전분립이 거의 관찰되지 않았으나, 미토콘드리아, 골지체 등 다양한 세포소기관들이 다시 관찰되었다. 한편, scopolamine과 hyoscyamine 함량은 배양 60일부터 다소 증가하다가 이후 90일까지 급격히 증가하였고, 이후에는 급격한 감소를 보였다. 본 연구 결과 tropane alkaloid 생합성은 탈분화 및 분화 과정에 영향을 받으며, 세포 내 전분립 생성은 기관분화와 더불어 물질생합성과 매우 밀접한 관련성이 있을 것으로 추정된다.

Plasma glutathione peroxidase (pGPx) is an extracellular antioxidative selenoenzyme which has been detected in various adult tissues, but little is known about the expression and distribution of pGPx during embryogenesis. To investigate the expression patterns of pGPx during embryogenesis, we performed quantitative real-time PCR, in situ hybridization, Western blot, and immunohistochemistry analyses in whole embryos or each developing organ of mice on embryonic days (E)7.5–18.5. In whole embryos of E7.5–8.5, pGPx mRNA was more typically expressed in extra-embryonic tissues including ectoplacental cone, trophectoderm, and decidual cells than in embryos. However, after E9.5, pGPx mRNA and protein levels were increased in the embryos with differentiation and growth, but trended to gradually decrease in the extra-embryonic tissues until E18.5. In sectioned embryonic tissues on E13.5–18.5, pGPx mRNA and protein were mainly expressed in the developing nervous tissues, the sensory organs, and the epithelia of lung, skin, and intestine, the heart and artery, and the kidney. In particular, pGPx immunoreactivity was very strong in the developing liver. These results indicate that pGPx is spatio-temporally expressed in various embryonic organs as well as extra-embryonic tissues, suggesting that pGPx may function to protect the embryos against endogenous and exogenous reactive oxygen species during organogenesis.

Zinc oxide nanoparticles (nZnO) are used in a various range, including ceramic manufacture, photocatalysis, UV filters, and the food industry. However, little is known about the effects of micro- and nano-particles during mouse embryo organogenesis. To determine whether ZnO affects size-dependent anomalies during embryonic organogenesis, mouse embryos were cultured for two days with 300 ug/ml micro ZnO (mZnO;80±25 μm) and nZnO (< 100 nm) and the developmental changes were then investigated. Quantity of Zn by inductively coupled plasma mass spectrometry analysis, and expression patterns of various antioxidant enzymes in the embryos were investigated. Embryos exposed to mZnO or nZnO exhibited severe retardation of growth and development. In embryos exposed to mZnO and nZnO, yolk sac diameter, crown-rump length, and head length were significantly diminished. The morphological parameters, including yolk sac circulation, allantois, flexion, heart, hindbrain, midbrain, forebrain, otic system, optic system, branchial bars, maxillary process, mandibular process, olfactory system, caudal neural tube, forelimb, hindlimb, and somites in mZnO and nZnO-treated groups were significantly decreased. Zn absorption of the nZnO-treated group was significantly higher than that of the mZnO-treated group. Significantly decreased levels of CuZn-SOD, Mn-SOD, cGPx, and PHGPx mRNA were observed in the ZnO-treated group. In addition, antioxidant enzyme mRNA expressions of the nZnO group were significantly diminished, less than those of the mZnO treated group. These findings indicate that 300 ug/ml ZnO showed abnormality and nZnO may have a more severe effect than mZnO in developing embryos.

Platycodon grandiflorum (Bell flower) is an important plant that has traditionally been used as herbal medicine for the treatment of cough, phlegm, sore throats, lung abscesses, chest pains, dysuria, and dysentery. The present study was initiated to investigate the feasibility of inducing shoot and root organogenesis in cultured explants of P. grandiflorum in a range of culture media and through use of various plant growth regulators (PGRs). The plantlets (Stem containing one node) were isolated and cultured on different concentrations of Murashige and Skoog (MS) medium supplemented with PGRs. We found that proliferation and elongation of shoots and roots could be achieved on ¼ MS for P. grandiflorum with wild and green petals and on ⅛ MS for P. grandiflorum with double petals. The highest levels of development and elongation of adventitious shoots and roots were observed when petal explants were cultured on ¼ MS (pH 3.8) supplemented with 5% sucrose. Increasing the agar concentration reduced shoot growth and rooting potential; nevertheless, the highest number of shoots and roots was observed on 0.6% agar. In the case of growth regulators, ¼ MS supplemented with 1 mg L-1 6-benzylaminopurine (BA) was found to be best for shooting, although higher concentrations of BA tended to reduce shoot and root elongation. The highest number of shoots was achieved on 0.5 mg ․ L-1 thidiazuron (TDZ) from double petal explants grown on ⅛ MS. However, root and shoot elongation were found to decrease when TDZ concentrations were increased. Low concentrations of kinetin, naphthalene acetic acid, indole acetic acid, and 3-indole butyric acid induced shoot and root proliferation and elongation. Taken together, our study showed that low concentrations of PGRs induced the greatest root formation and elongation, showing that the optimal concentration of PGRs for shoot proliferation was species-dependent.

Camptotheca acuminata, a native of South China is a well known natural source of monoterpene-indole alkaloid camptothecin(CPT), one of the most promising anti-tumoural compounds. This study was conducted to optimize plant growth regulators and culture conditions on plantlets regeneration through organogenesis from callus of Camptotheca acuminta. Callus were induced from various explants of in vitro germinated plantlets of C. acuminta using WPM medium containing 0.2 ㎎/L 2,4-D. Hypocotyl segments were exhibited higher embryogenic callus than the other explants. Shoot buds formation from embryogenic callus was affected by plant growth regulators, pre-treated dark condition and liquid culture. Organogenesis was optimal in WPM liquid medium containing 0.5 ㎎/L BA. The dark pre-treatment for 2 weeks before the solid culture was effective for organogenesis. The regenerated shoots were rooted in WPM medium with 0.2 ㎎/L NAA and successfully acclimated in green-house conditions.

한국의 전통적인 민간 약재로 사용되어져온 흰민들레의 잎절편체로 부터 기관형성을 통한 재분화를 시도하였다. 잎 절편체를 BA (0-4mg/L)와 2,4-D (0-1mg/L)가 혼합 첨가된 MS 기본 배지에 치상하여 배양한 결과, 배양 3주후부터 신초가 발생되었다. 신초 분화율은 2mg/L BA 첨가된 조건에서 가장 높았으며, 배양 4주째 4mg/L BA 첨가된 조건에서는 캘러스가 유기되었다. 잎령에 따른 부정아 형성율, 발생빈도, 건량, 생중량을 조사한 결과, 발아 후 7주된 잎 절편체에서 평균 11.5개의 신초를 형성하였다. 유도된 신초는 0.5mg/L NAA가 첨가된 배지에서 발근되었으며, 화분에 옮겨 완전한 식물체로 재분화 되었다.

A method for plant regeneration via organogenesis from Pelagonium inquinans leaf disc has been developed. Mature leaf explants were collected from field-grown plants and used for the induction of adventitious shoot regeneration on Murashige and Skoog (MS) medium supplemented with 3% (w/v) sucrose plus plant growth regulators. Maximum shoot organogenesis, with 11.8±1.5 shoots (98.6%) per leaf disc, was obtained with 2 mg/l N6-benzyladenine (BA) and 0.5 mg/l α-naphthyleneacetic acid (NAA) in 30 days. For rooting, the in vitro proliferated and elongated shoots were excised into 1.5-2 cm in length microcutting, which were plated individually on an half-strength MS (1/2MS) medium supplemented with 2% (w/v) sucrose plus various concentrations of indole-3-butyric acid (IBA). Shoots rooted with a frequency of 100% following culture on 1/2MS medium containing 0.5 mg/l IBA.

An efficient plant regeneration of C. asiatica was achieved from organogenesis using petiole explants of in vitro plantlet on MS basal medium controled with different plant growth regulators (NAA,2,4-D, IAA kinetin, and BA). Best results that 50%, efficiency of regeneration per explant for regeneration were obtained with IAA 17.13 μM and BA 8.9 μM. Formation of adventitious shoots via organogenesis from the petiole explant was verified by histological sectioning of plantlets. Regenerated plants were transplanted into soil.

약용식물로 쓰이는 선피막이 (Hydrocotyle maritima Honda)를 기내에서 재분화 가능성에 대해 조사하였다. 선피막이의 엽병 절편체를 식물생장조절제 (0~5 mg/l, NAA와 0~5 mg/l 2,4-D)가 단독 또는 0.1~2 mg/l BA와 조합 첨가된 배지 에서 6주동안 배양하였다. 2,4-D 또는 NAA을 단독으로 처리해도 캘러스가 발생하나, 가장 좋은 결과는 각각 0~5 mg/l BA, 5 mg/l NAA와 0.5 mg/l BA조합구에서 나타났다. Kinetin를 3 mg/l로 처리시 캘러스로부터 가장 많은 신초 (캘러스당 12개)을 발생시켰다. 또한, 0.5 mg/l 2,4-D와 0.5 mg/l BA가 첨가된 배지에서 유도된 배 발생 캘러스를 호르몬이 없는 배지에서 배양하였을 때, 체세포 배가 분화되었고, 식물체로 더 잘 발달하였다.