Dong Hwi Kim

Optimal Harvesting Time of Ginseng Seeds and Effect of Gibberellic Acid (GA3) Treatment for improving Stratification Rate of Ginseng (Panax ginseng C. A. Meyer) Seeds

This study was performed to identify optimal harvesting time of ginseng seeds and to examine the effect of GA3 treatment for improvement of seed stratification rate. Ginseng seeds harvested from Land race, Chunpoong and Yun- poong cultivar in July 20 were tested for stratification rate. It was shown that stratification rates of land race, Yunpoong and Chunpoong cultivar were 94.1%, 93.1%, and 82.6%, respectively. Seeds of Chunpoong cultivar harvested 10-15 days later showed a comparable stratification rate to that of Land race, indicating that late harvest of Chunpoong seeds is beneficial for the increase of stratification rate. The higher stratification rate was found in mature seeds (92.3%) than immature seeds (37.8%), both of which were harvested in July 20. Stratification rate of mature seeds harvested in July 15 was 87.5%, dem- onstrating optimal harvesting time of ginseng seeds with higher stratification rate is after mid-July. An exponential growth of endosperms of ginseng seeds was observed from early June to mid-June and then slow growth was observed. There was no obvious growth of embryos from fertilization to mid-August. After the this time, embryos quickly grew until late October. Thus, appropriate stratification control is essential during the period (from early September to late October) in order to optimize embryo growth and development. While no increase of stratification rate was observed in seeds treated with 50 ppm of GA3, significant increases were observed in seeds treated with 100 ppm of GA3. At this concentration of GA3, the stratification rate of Land race, Chunpoong and Yunpoong cultivar was 95.0%, 95.3%, and 96.5%, respectively.

Analysis of Mitochondrial DNA Sequence and Molecular Marker Development for Identification of Panax Species

This study describes the identification of Panax species using mitochondrial consensus primers. Initially, a total of thirty primers were tested in ten Korean ginseng cultivars and two foreign Panax species, P. quinquefolius and P. notoginseng. In the polymerase chain reaction (PCR) amplification results, three primers (cox1, nad1/2-3 and nad2/1-2) generated co-dominant polymorphic banding patterns discriminating Korean ginseng cultivars from P. quinquefolius and P. notoginseng. However, these primers could not generated polymorphisms among the Korean ginseng cultivars, and simply represented species-specific polymorphisms for P. quinquefolius and P. notoginseng. Primers PQ91 and PN418 were designed from the consensus sequence of nad1/2-3 region. Two banding patterns (A or B) were detected in PQ91. Korean ginseng cultivars and P. notoginseng shared the same banding pattern (A type) and P. quinquefolius was identified another banding pattern (B type). In the case of PN418, two banding patterns (A or B) were detected in the Korean ginseng cultivars and two foreign Panax species. Korean ginseng cultivars and P. quinquefolius shared the same banding pattern (A type) and P. notoginseng was identified another banding pattern (B type). The combination banding patterns of three Panax species, Korean ginseng cultivars (Panax ginseng C. A. Mey.), P. quinquefolius and P. notoginseng, was identified as `AA`, `BA` and `AB`, respectively. Consequently, PQ91 and PN418 primer sets can be used to distinguish among Panax species.

Effects of Salt in Soil Condition on Chlorophyll Fluorescence and Physiological Disorder in Panax ginseng C. A. Meyer

Background : Excessively high concentration of sodium ion causednutrient deficiency and significantly decrease growth. This study was carried out to determine the limiting concentration range of sodium ion in the soil of ginseng field. Methods and Results : The growth of the ginseng cultivar Chunpoong reduced with increase in salinity, and the rate of growth reduction was higher in shoots than that of roots. Particularly, ginseng plants cultivated at high level of nitrate nitrogen or sodium may suffer delayed development and stunted growth. Chlorophyll damage occurred on the leaves of ginseng planted in relatively high levels (> ) of sodium ion, as determined by the fluorescence reaction. The incidence of physiological disorder in ginseng cultivated at 249 sites was correlated with the concentration of sodium ion in the soils. About 74% of ginseng fields in which physiological disorders occurred had concentrations of sodium ion in soil greater than . In contrast, the concentration of sodium ions at 51 of 85 sites where no damage occurred was relatively (). Conclusions : The concentration of sodium ion in soil of ginseng fields can be classified into three levels optimum (), permissible allowance (0.15 - 0.2) and excessive (> 0.2).

Applications of molecular markers in the discrimination of Panax species and Korean ginseng cultivars (Panax ginseng)

The development of molecular markers is one of the most useful methods for molecular breeding and marker-based molecular associated selections. Even though there is less information on the reference genome, molecular markers are indispensable tools for determination of genetic variation and identification of species with high levels of accuracy and reproducibility. The demand for molecular approaches for marker-based breeding and genetic discriminations in Panax species has greatly increased in recent times and has been successfully applied for various purposes. However, owing to the existence of diverse molecular techniques and differences in their principles and applications, there should be careful consideration while selecting appropriate marker types. In this review, we outline the recent status of different molecular marker applications in ginseng research and industrial fields. In addition, we discuss the basic principles, requirements, and advantages and disadvantages of the most widely used molecular markers, including restriction fragment length polymorphism, random amplified polymorphic DNA, sequence tag sites, simple sequence repeats, and single nucleotide polymorphisms.

High frequency somatic embryogenesis and plant regeneration of interspecific ginseng hybrid between Panax ginseng and Panax quinquefolius

Background Interspecific ginseng hybrid, Panax ginseng × Panax quenquifolius (Pgq) has vigorous growth and produces larger roots than its parents. However, F1 progenies are complete male sterile. Plant tissue culture technology can circumvent the issue and propagate the hybrid. Methods Murashige and Skoog (MS) medium with different concentrations (0, 2, 4, and 6 mg/L) of 2,4-dichlorophenoxyacetic acid (2,4-D) was used for callus induction and somatic embryogenesis (SE). The embryos, after culturing on GA3 supplemented medium, were transferred to hormone free ½ Schenk and Hildebrandt (SH) medium. The developed taproots with dormant buds were treated with GA3 to break the bud dormancy, and transferred to soil. Hybrid Pgq plants were verified by random amplified polymorphic DNA (RAPD) and inter simple sequence repeat (ISSR) analyses and by LC-IT-TOF-MS. Results We conducted a comparative study of somatic embryogenesis (SE) in Pgq and its parents, and attempted to establish the soil transfer of in vitro propagated Pgq tap roots. The Pgq explants showed higher rate of embryogenesis (~56% at 2 mg/L 2,4-D concentration) as well as higher number of embryos per explants (~7 at the same 2,4-D concentration) compared to its either parents. The germinated embryos, after culturing on GA3 supplemented medium, were transferred to hormone free ½ SH medium to support the continued growth and kept until nutrient depletion induced senescence (NuDIS) of leaf defoliation occurred (4 months). By that time, thickened tap roots with well-developed lateral roots and dormant buds were obtained. All Pgq tap roots pretreated with 20 mg/L GA3 for at least a week produced new shoots after soil transfer. We selected the discriminatory RAPD and ISSR markers to find the interspecific ginseng hybrid among its parents. The F1 hybrid (Pgq) contained species specific 2 ginsenosides (ginsenoside Rf in P. ginseng and pseudoginsenosides F11 in P. quinquefolius), and higher amount of other ginsenosides than its parents. Conclusion Micropropagation of interspecific hybrid ginseng can give an opportunity for continuous production of plants.

In vitro grown thickened taproots, a new type of soil transplanting source in Panax ginseng

Background The low survival rate of in vitro regenerated Panax ginseng plantlets after transfer to soil is the main obstacle for their successful micropropagation and molecular breeding. In most cases, young plantlets converted from somatic embryos are transferred to soil. Methods In vitro thickened taproots, which were produced after prolonged culture of ginseng plantlets, were transferred to soil. Results Taproot thickening of plantlets occurred near hypocotyl and primary roots. Elevated concentration of sucrose in the medium stimulated the root thickening of plantlets. Senescence of shoots occurred following the prolonged culture of plantlets. Once the leaves of plantlets senesced, the buds on taproots developed a dormant tendency. Gibberellic acid treatment was required for dormancy breaking of the buds. Analysis of endogenous abscisic acid revealed that the content of abscisic acid in taproots with senescent shoots was comparatively higher than that of taproots with green shoots. Thickened taproots were transferred to soil, followed by exposure to gibberellic acid or a cold temperature of 2°C for 4 mo. Cold treatment of roots at 2°C for 4 mo resulted in bud sprouting in 84% of roots. Spraying of 100 mg/L gibberellic acid also induced the bud sprouting in 81% roots. Conclusion Soil transfer of dormant taproots of P. ginseng has advantages since they do not require an acclimatization procedure, humidity control of plants, and photoautotrophic growth, and a high soil survival rate was attained.

Internal Transcribed Spacer Barcoding DNA Region Coupled with High Resolution Melting Analysis for Authentication of Panax Species

Background : Correct identification of Panax species is important to ensure food quality, safety, authenticity and health for consumers. This paper describes a high resolution melting (HRM) analysis based method using internal transcribed spacer (ITS) and 5.8S ribosomal DNA barcoding regions as target (Bar-HRM) to obtain barcoding information for the major Panax species and to identify the origin of ginseng plant. Methods and Results : A PCR-based approach, Bar-HRM was developed to discriminate among Panax species. In this study, the ITS1, ITS2, and 5.8S rDNA genes were targeted for testing, since these have been identified as suitable genes for use in the identification of Panax species. The HRM analysis generated cluster patterns that were specific and sensitive enough to detect small sequence differences among the tested Panax species. Conclusion : The results of this study show that the HRM curve analysis of the ITS regions and 5.8S rDNA sequences is a simple, quick, and reproducible method. It can simultaneously identify three Panax species and screen for variants. Thus, ITS1HRM and 5.8SHRM primer sets can be used to distinguish among Panax species.

Effects of GA3 and Alternating Temperature on Breaking Bud Dormancy of Panax ginseng C. A. Meyer Seedling

Background : Considerable time and effort is required to develop new Panax ginseng varieties. Ginseng breeders have been developing techniques to shorten the breeding cycle to resolve this problem. In this study, we investigated the effects of adding GA3 and alternating temperature (AT, 2℃→ −2℃ → 2℃) on breaking bud dormancy in the varieties (Chungsun and Sunun) of ginseng root. Methods and Results : The GA3 soaking treatment and AT were applied to one year old roots, which greatly accelerated the emergence of new buds. In one year old roots, new buds emerged from the 4th day post transplanting and after breaking dormancy with GA3 and AT treatments. The emergence of new buds was completed within two weeks. The rate of bud emergence for Chungsun was 60% - 98% over 15 - 60 days after the AT and GA3 treatments. The emergence rate of Sunun was 46% - 92%. Normal growth of the ginseng seedling was observed in spite of the early breaking of bud dormancy by combined GA3 and AT treatments. Conclusions : GA3 and AT treatments shortened the dormancy period and facilitated the stable emergence of ginseng seedlings. However, some plants suffered deformities and early sprouting owing to the combined GA3 and AT treatments. Early sprouting was free from dormancy after leaf fall from the of aerial part of the plant.

Clinical Study on Food Safety evaluation of Panax ginseng

*Department of Herbal Crop Research, NIHHS, RDA, Eumseong 369-873, Korea.**College of Chinese Medicinal Materials, Jilin Agricultural University, Changchun 130118, China.ABSTRACT : In this study, the clinical safety and toxicology of oral ingestion of supplement capsules containing ginsengradix was investigated in healthy young volunteers. This study was a pilot randomized, double blinded, placebo controlledtrial. The healthy volunteers were divided into 6 groups of 20 each (10 males and 10 females). They took the ginseng powderfor 35 days (3g/day) for safety evaluation. There were measured general healthy levels such as hematological, biochemical andelectrocardiographic parameters. After the first week, besides Korean white ginseng the other treatments led to an significantincrease of white blood cells. Korean red ginseng increased UREA (blood urea nitrogen) in healthy volunteers, but it didn’texceed the range of normal values, and in the subsequent process of treatment there is no effect of elevating UREA. After thethree weeks, Korean white ginseng showed relatively low the content of blood glucose and low-density lipoprotein cholesterol.After the five weeks, compared with the other treatments, Korean red ginseng increased white blood cells, platelet distributionwidth and average volume of platelet. Korean white ginseng decreased low-density lipoprotein cholesterol. American ginsengdecreased blood creatinine in healthy volunteers. In conclusion, through test the blood routine, urine routine, liver function,renal function, blood glucose, blood lipid and electrocardiogram, the healthy volunteers continuous taking ginsengfor 35 days (3 g/day) is safe and reliable, and have no obvious adverse reactions and side effects.Key Words : Panax ginseng, Panax quinquefolium, Food Safety

Difference in Growth Characteristics of 5-Year-Old Ginseng Grown by Direct Seeding and Transplanting

Background : In order to determine the effects of planting methods on root growth of ginseng varieties, two different methods, direct seed sowing and transplanting were compared in terms of their effects on different root growth characteristics. Methods and Results : Higher fresh root weight was observed in ginseng grown by direct seed sowing. Direct seed sowing of three cultivars (Sunhyang, Chungsun and K-1) resulted in higher yield, whereas no difference was observed in the yield of one cultivar (Chungsun). Gumpoong was highly tolerant to physiological stress, as it showed fewer symptoms of rusty and rough skin root diseases in both direct seed sowing and transplanting. The average main root length per total root length of ginseng grown by direct seed sowing was 33.6%, whereas that of ginseng grown by the average of those by transplanting was 22.4%. Other root growth characteristics, including root length, main root diameter, and number of side roots, improved when the direct seed sowing method was used. Conclusions : To our knowledge, this is the first study reporting the differences in root growth parameters of ginseng varieties grown by direct seed sowing or transplanting at the same planting density. Because of the advantages of direct sowing during ginseng planting, developing new varieties and improving cultivation methods are imperative.