내염성 유전자의 식물형질전환용 벡터개발 및 현사시의 형질전환

문흥규,노은운,이재순 한국산림과학회 1998 한국임학회 학술발표논문집 Vol.1998 No.-

희귀 및 멸종위기 식물 덩굴용담의 기내생장에 미치는 광질 및 환기효과

문흥규,박소영 한국식물생명공학회 2008 식물생명공학회지 Vol.35 No.3

Effects of light generated by LEDs on shoot growth and rooting of Tsuru-rindo (Tripterospermum japonicum) were evaluated. Apical shoots (one or two node with 3-4 leaves) were cultured on MS basal medium with 3% sucrose and maintained for four weeks under five different light qualities: fluorescent lamp (F), 100% red LED (R), 70% red LED+30% blue LED (R7B3), 50% red LED+50% blue (R5B5), or 100% blue LED (B). Rooting was promoted by both red light and fluorescent lamp, and the effect was further promoted under the ventilation. Red light enhanced shoot node elongation, whereas blue light appeared to suppress it. Growth of shoots and leaves were enhanced under the ventilation irrespective of the different light qualities. Under the ventilated condition, total fresh weight of plants was highest in R7B3 LED as 257.7 mg per plant. Dry matters, which are used for index of plant growth, were lowest under red light, whereas it was highest under blue light. The dry matter was inclined to getting higher by ascending the ratio of blue light and red light. Total chlorophyll content was highest in both R7B3 LED and R5B5 LED under ventilation as 29.5 and 31.2, respectively. Above results suggest that light quality optimization could be an important factor to foster in vitro growth of the species. Ventilation treatment appeared to be another important factor to induce normal shoot growth and rooting

음나무 배발생 캘러스의 증식 및 체세포배 발달을 위한 액체 현탁 배양조건 확립

문흥규,김선자 한국식물생명공학회 2009 식물생명공학회지 Vol.36 No.1

This study was conducted to establish the optimal suspension culture system for both the propagation of embryogenic cells (ECs) and the induction of somatic embryos (SEs) of Kalopanax septemlobus. The proliferation rate of ECs was reduced as the inoculum density was increased; the highest rate was obtained when 0.1 g/100 ml of cells was initially inoculated. According to the analysis of cell growth pattern and cell growth cycle (G1, S and G2/M), the cell growth started in 5 days culture initiation, grew rapidly until 15 days and then decreased gradually. Distinctive changes of the cell growth cycle by the culture periods was also observed; the growth cycle was doubled from initial 5.6% to 11.7% of S stage in 5 days culture and then reached in stable stages again. Therefore, the results indicated that a 15-day-cycle was the optimal culture period for the propagation of the ECs through the suspension culture. Furthermore, the cell inoculum density was also important for the induction of SE; more than 65% of SEs at the torpedo stage was induced by using the low level of cell inoculum (0.5 g/L), while the higher inoculum densities were rapidly reduced the proportion of SEs at that stage. Although the higher inoculum density delayed the development of SE, it did not affect the proportion of SEs at the globular and heart stage. In conclusion, this study showed that the suspension culture of the Kalopanax septemlobus ECs through the control of inoculum density was an efficient way for both the propagation of ECs and the induction of SEs, suggesting that the development of this system might help to reduce the culture period for the somatic embryo production. 본 실험은 음나무 배발생 세포의 증식 및 체세포배 발달을 위한 액체 현탁 배양조건의 확립을 위해 수행되었다. 배발생 세포의 생 장율은 접종 밀도가 증가할수록 감소하였다. 배발생 세포의 증식 에 가장 효과적인 접종 밀도는 0.1 g/100 ml 로서 이 농도에서 세포 의 생장율이 가장 높았다. 배양기간에 따른 배발생 세포의 생장 패 턴 및 세포 주기 (G1, S, G2/M) 분석 결과, 세포의 생장은 배양 5일 후부터 증가가 시작되어 15일 까지 급격히 생장하였으며 그 이후 에는 점차 감소하였다. 배양 기간 별 세포 주기 (cell cycle) 의 변화 가 명확하게 관찰되어 배양 5일째 S기는 초기의 5.6% 에서 11.7% 로 두 배 정도 증가하였으며, 배양 15일 이후부터는 다시 초기의 세포 주기로 되돌아 가면서 안정화되는 것으로 나타났다. 따라서 음나무 배발생 세포의 현탁배양은 15일의 주기로 배양하는 것이 증식에 가장 효과적인 것으로 생각되었다. 한편 배발생 세포에서 체세포배의 유도는 배양초기의 접종 밀도가 중요한 것으로 나타났 다. 0.5 g/L 의 낮은 밀도로 접종 시에는 65% 이상의 어뢰형 배가 유도된 반면 접종 밀도가 높아질수록 어뢰형으로의 배발달은 급격 히 감소하였다. 초기의 접종 밀도가 증가할수록 특히 어뢰형 배의 발달은 지연되었으나 구형 및 심장형 배의 유도는 초기 접종밀도 에 영향을 받지 않았다. 이상의 실험결과로 음나무 액체 배양 시 초기 접종 밀도를 조절함으로써 배발생 캘러스의 증식 및 체세포 배를 효과적으로 유도할 수 있었으며 이는 체세포배 생산을 위한 배양 기간의 단축이 가능함을 보여주는 결과이다.

멸종위기 식물 덩굴용담의 기내번식에 미치는 생장조절제 효과

문흥규,김선자,박소영,김용욱,이재선 한국식물생명공학회 2009 식물생명공학회지 Vol.36 No.1

Various plant growth regulators were tested for shoot proliferation of Tripterospermum japonicum, a rare and endangered species. Among the six different media tested, MS medium was the best for the shoot growth. Whereas BA, upto 3 mg/L, significantly increased shoot proliferation rate, it suppressed the rate at higher levels. Neither kinetin nor TDZ was so effective in proliferating shoots as BA. As for rooting, TDZ strongly inhibited it even at very low concentration though spontaneous rooting was frequently observed from the proliferated shoots during culture of lower concentration BA or kinetin. In contrast, shoot elongation was significantly promoted by GA3. More than 90% of the proliferated plantlets could be transplanted via cuttings into pots containing artificial soil mixture where they rooted and resumed normal growth. Most of the plants bloomed to bear fruits in the following year. 멸종위기 식물 덩굴용담의 기내증식법 개발을 위하여 기내증식 에 미치는 생장조절제의 효과를 시험하였다. 6가지의 기본배지 시 험에서 MS 배지가 줄기증식에 가장 적합한 것으로 나타났다. 싸이 토키닌 가운데 BA 처리는 3.0 mg/L 농도까지 증식에 유의적인 증 식효과를 보였고, 그 이상 농도에서는 증식이 억제되었다. Kinetin 과 TDZ는 BA 만큼 증식에 효과적이지 못했다. 발근에 있어 TDZ 는 저농도에서도 발근을 현저히 억제하였으나, BA와 kinetin은 저 농도 처리 조건에서 자발적인 발근이 이루어 졌다. 한편 지베렐릭 산 (GA3)의 처리는 줄기신장의 유의적인 효과가 있었다. 증식중인 줄기는 인공상토에 기외삽목하여 90% 이상 발근이 가능하고 정상 적인 생장이 이루어졌다. 대부분의 식물체는 토양이식 다음해에 개화되고 과실을 맺었다.

상수리나무 기내 (器內) Axillary Bud 의 치상부위에따른 (置床部位) 다경 (多莖) 및 발근유도 (發根誘道) 효과

문흥규,김재헌,박재인 ( Heung Kyu Moon,Jae Hun Kim,Jae In Park ) 한국산림과학회 1987 한국산림과학회지 Vol.76 No.4

This study was conducted to examine the position effect of axillary buds on shoot multiplication and rooting with 1-year-old seedlings of Quercus acutissima. Shoot multiplication was greatly affected by axillary bud position: Productivity of multiple shoots were decreased in the order of top, mid and basal explants respectively. The best shoot multiplication (mean 6.1 shoots per explant) was obtained on WPM medium containing 1.0㎎/Q BAP and 0.1㎎/Q NAA using basal explant after 4-week-culture. Rooting was also greatly influenced by position. Its percentage was increased in the order of top, mid and basal explant respectively. Root initiation was better and more rapid on ½MS medium than GD medium. High rooting percentage (100%) was obtained on ½MS medium containing 0.2㎎/ℓ IBA after 15 days culture. Sucrose concentrations did not effect on rooting. However root development and shoot growth were greatly affected by them. Root was developed shortly on 1-2% levels and shoot growth was getting retarded, whereas both of them did not show significant difference at 3-6% levels. Rotting was decreased on 7-8% levels gradually, but shoot and leaf condition was better than any other concentrations. Survival rate of rooted explants in pot was varied according to the position of explants. Seedlings of top part were survived up to about 50% but most of mid and basal part seedlings did not survive over 4 weeks even in high humidity condition. Seedlings in pots showed normal growth over 10 months but most of them showed the condition of premature leaf shedding.

희귀 수종 산개나리의 기내 번식

문흥규,석진영,김선창 ( Heung Kyu Moon,Gene Young Suk,Sun Chang Kim ) 한국산림과학회 1997 한국산림과학회지 Vol.86 No.4

Axillary bud explants from 3-year-old seedlings of Forsythia saxatilis N., rare and endangered species in Korea, were cultured on Murashige and Skoog`s medium. The effect of various cytokinins (BAP, kinetin, and zeatin) at the different concentration(0.2, 0.5 and 1.0㎎/ℓ) was tested. Although an apparent shoot proliferation was not observed, zeatin showed slight promotional effect on normal shoot and leaf development. Both shoots and adventive roots could be induced simultaneously when the explants were cultured on the medium with kinetin, but adventive rooting was gradually reduced according as BAP and zeatin concentrations increased. Axillary shoot growth was promoted by the etiolation treatment. Shoot proliferation has been maintained more than three years with consecutive subculture. Rooted plantlets were successfully acclimatized in the artificial soil mixture and showed normal growth after transplantation into field.

두릅(Aralia elata)의 체세포배 유도, 발아 및 식물체 재분화에 미치는 요인

문흥규,오경은,손성호 한국식물생명공학회 1999 식물생명공학회지 Vol.26 No.4

두릅나무 (Aralia elata Seem.) 2년 생 11개체의 동아를 1.0 mg/L 2,4-D, 3% sucrose, 0.3% gelrite를 포함한 MS 고체 배지에서 체세포배 형성, 발아 및 식물체 재분화에 미치는 개체목의 효과를 조사하였다. 배발생 캘러스의 유도는 개체목에 따른 차이가 있었으나 발아 및 식물체 재분화는 차이가 뚜렷하지 않았다. 배발생 캘러스는 대체로 노란색을 띠었고, 조성은 비배발생 캘러스보다 부드럽고 생장은 다소 느렸다. 체세포배의 발아 및 식물체 재분화는 MS배지의 염류 농도가 크게 영향하였고, 기본배지보다는 염류 농도를 반감시킨 1/2 MS배지에서 BA및 ABA의 처리에 관계없이 양호한 배발아 및 식물체 재분화를 나타냈다. BA처리는 0.1mg/L 농도에서 배축과 자엽 발달에 다소 촉진효과를 나타냈으나 발아와 재분화율은 기본배지보다 우수하지 못했다. 발아되는 배는 자엽수 및 형태에 있어 다양한 변이를 나타냈다. ABA처리는 두릅 체세포배의 발아 및 재분화에 비효과적이었다. 재분화된 식물체는 인공배양토에서 95%이상 환경순화되어 포지에 이식 후 형태적 변이 없이 정상생장이 가능하였다. In order to find optimum conditions for somatic embryogenesis from different individual (2-year-old) in Aralia elata were cultured on MS medium supplemented with 1.0 mg/L 2,4-D, 3% sucrose, and 0.3% gelrite. We also investigated the effect of MS medium salt concentration, BA and ABA on the embryo germination and plant regeneration. While noticeable difference was observed on somatic embryo induction among different individual tree, no apparent difference was seen in both germination and regeneration frequencies. Compared with nonembryogenic calli, embryogenic calli tended to look yellow and/or pale brown in color, slowly growing and soft in their texture. Regardless of BA or ABA treatment, half-strength MS salt medium proved to be better than full strength MS medium in both embryo germination and plant regeneration. Both hypocotyl and cotyledon developments were slightly promoted by adding 0.1 mg/L BA. However, its effect on germination and regeneration seemed inferior to control. ABA treatment on somatic embryos at their torpedo and early cotyledonary stages resulted in poor response in germination and regeneration. Although most regenerated plantlets varied greatly in cotyledon number and shape, they could be developed into normal plants after 4 weeks in culture. More than 95% plantlets were acclimatized in an artificial soil mixture, successfully transplanted to nursery bed and grew normally without any phenotypic abnormalty.

두릅나무 체세포배 유래 소식물체의 순화에 미치는 배양토 및 공급액의 효과

문흥규,배찬호,김용욱,이재순,이재선 한국식물생명공학회 2001 식물생명공학회지 Vol.28 No.5

두릅나무의 체세포배 유래 소식물체의 효과적인 순화 방법을 개발하고자 플라스틱 용기를 사용 4가지 인공배양토 및 MS 액체배지 등 7가지 수용액 처리를 통해 활착률 및 생장을 조사하였다. 상토는 perlite가 vermiculite보다 활착 및 생장에 양호하였고, 입자가 클수록 활착률 및 생장이 좋았다. 액체배지는 2차 증류수 처리가 가장 양호하여 96%의 활착률을 나타냈으며, 염류농도를 1/4 및 1/8로 낮춘 MS배지는 각각 92%의 활착률을 보인 반면 기본배지 및 1/2 MS배지는 활착률이 저조하였다. 한편 0.1% hyponex와 2% sucrose 처리는 84% 및 76%의 활착률을 각각 나타냈다. 결론적으로 두릅나무의 체세포배 유래 소식물체의 순화는 입자가 큰 perlite를 사용하고 증류수 처리 혹은 염류 농도를 낮춘 MS배지 처리로 효과적인 순화가 가능함을 보여주었다. In order to develop effective acclimatization methods for Aralia elata plantlets regenerated from somatic embryos, various acclimatizing conditions were compared regarding both survival rate and growth of the plantlets. The plantlets were transplanted into plastic boxes containing artificial soil in the presence of either several levels of MS liquid media, distilled water, 2% sucrose or 0.1% hyponex solution. They were then cultured by spraying of distilled water twice a week and maintained in the normal tissue culture room. Perlite was proved to be better than vermiculite on survival rate and growth of the plantlets. As the size of perlite (larger than 0.2 cm in diameter) increased, both the survival rate and growth of the plantlets improved. Among the various MS liquid media and different aqueous solutions tested, distilled water appeared to result in the best survival rate and growth. MS media were also effective in increasing survival rate and supporting growth when diluted to 1/4 and/or 1/8. The acclimatized plantlets could be transplanted directly onto the nursery bed and grown normally. The above results suggest that plantlets regenerated from somatic embryos of Aralia elata be effectively acclimatized using a plastic box containing perlite with distilled water treatment.

두릅나무 15개체의 체세포배 유도 및 식물체 재분화에 미치는 유전자형의 효과

문흥규,홍용표,김용욱,이재순 한국식물생명공학회 2001 식물생명공학회지 Vol.28 No.3

Winter bud explants from 15 individual angelica tree (Aralia elata) were cultured in vitro to find out optimal conditions for somatic embryo induction as well as plant regeneration. Calli are induced and grown on MS medium supplemented with 1.0 mg/L 2,4-D for 4 weeks and subcultured on a half-strength MS medium without phytohormones to induce somatic embryos. Inter-simple sequence repeat (I-SSR) markers were analyzed with total DNAs extracted from the trees. Genotype effects on somatic embryo induction were examined by cluster analysis. Callus induction rate varied from 58.5 to 100% among the genotypes. Somatic embryo induction rate also greatly varied from 0 to 100% among the genotypes. There was a significant difference in somatic embryo induction rate even among the individual trees that showed close genetic relationships each other. This suggested that somatic embryo induction rate in Aralia elata be influenced by a few major specific genes rather than whole genomic similarity among individual trees. Four individuals of Ulneong-7, Cheju-1, Shingu and China, which are recalcitrant to somatic embryo induction, turned out to have a close genetic relationship, suggesting that both physiological and genetic factors affect somatic embryo induction. The results suggest that genotype selection be the most important factor to achieve an efficient propagation, although cultural optimization through medium and explant manipulation may also play crucial roles in somatic embryogensis as well as plant regeneration of these species.

액아배양에 의한 유묘 및 성숙 히어리나무의 기내변식

문흥규,노은운,하유미,심경구 한국식물생명공학회 2002 JOURNAL OF PLANT BIOTECHNOLOGY Vol.29 No.2