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다양한 계통의 옥수수 미성숙배로부터 캘러스 유도와 식물체 재분화
홍준기,박기진,이강섭,김둘이,김주곤,이승범,서은정,이연희,Hong, Joon Ki,Park, Ki Jin,Lee, Gang-Seob,Kim, Dool Yi,Kim, Ju-Kon,Lee, Seung Bum,Suh, Eun Jung,Lee, Yeon-Hee 한국식물생명공학회 2017 식물생명공학회지 Vol.44 No.1
옥수수의 최적 조직 배양 조건을 확립하기 위하여 옥수수 국내 5 계통과 국외 11 계통 총 16 계통을 포트와 포장 재배하여 미성숙 배를 분리하여 배발생 캘러스 유도 및 식물체 재분화율을 조사하였다. MS 배지에 auxin으로 1.5 mg/L Dicamba와 0.5 mg/L 2,4-D 가 첨가된 배지에서 캘러스 형성은 본 실험에 사용된 옥수수 계통 모두에서 높은 빈도로 유도되었으며, 캘러스로부터 식물체 재분화는 5mg/L zeatin이 첨가된 재분화 배지에서 높은 재분화율을 보였다. 또한 포장에서 재배된 옥수수로부터 미성숙 배를 분리하여 사용하였을 때 캘러스 유기 및 식물체 재분화 효율이 높았던 것으로 보아 미성숙 배를 분리하기 위한 옥수수 상태 및 genotype이 중요한 영향을 준다는 것을 알 수 있었다. 본 실험을 통하여 배 발생 캘러스 형성 및 식물체 재분화 효율이 조사된 옥수수 계통들은 생명공학 기술을 활용한 신품종 개발을 위한 형질전환 시스템 개발에 유전자원으로 활용될 수 있는 정보를 제공할 것으로 사료된다. We investigated the callus induction and plant regeneration ability of 16 maize genotypes, including the Korean inbred lines, using 9 to 15 day-old immature zygotic embryos from maize grown in pots and from field cultures. Immature zygotic embryos placed on MS medium supplemented with L-proline 0.7 g/L, MES 0.5 g/L, Dicamba 1.5 mg/L, 2,4-D 0.5 mg/L, $AgNO_3$ 4 mg/L, and sucrose 20 g/L, showed the highest frequency of callus induction. The highest number of shoots regenerated when the embryogenic callus were transferred to MS medium supplemented with 5 mg/L zeatin. The root formation was observed when shoots were grown on MS medium supplemented with 0.2 mg/L indole-3-butyric acid (IBA). Additionally, under the same culture conditions, immature zygotic embryos from maize grown in the field also had a high frequency of plant regeneration. Except one genotype, 15 genotypes showed callus induction and shoot regeneration. Among the 16 genotypes tested, H99, B98, HW3, and B73 yielded the best plant regeneration. H99 showed maximum shoot formation from the primary embryogenic callus. The results suggest that genotypes and growth conditions of the maize plant plays very important roles for enhancing the embryogenesis competence of immature zygotic embryos. The successful regeneration from immature zygotic embryos of maize inbred lines provides a basis for molecular breeding of new cultivars by genetic transformation.
신윤철(Yun Cheol Shin),김주곤(Ju Kon Kim),남백희(Baek Hie Nahm) 한국응용생명화학회 1993 Applied Biological Chemistry (Appl Biol Chem) Vol.36 No.6
To characterize glutelins, the most abundant storage protein in rice, 13 complete coding sequences of glutelin genes from the database were analyzed. According to the phylogenic analysis, these genes could be classified into 5 groups, Group I to V. The degrees of homology were calculated to be in the range of 90 to 60%, but the patterns of hydrophobicity were similar in all the groups. Also, each group was found to have similar amino acid composition with variations in lysine content from 2.5 to 3.6% due to the point mutation of arginine to lysine. The isoelectric points of mature proteins and their basic chains of all the groups showed the value of about 9.0 and 10.0, respectively, while the isoelectric points of acidic chains in these groups showed the distinct value of 6.6, 6.7, 7.2, 8.4 and 7.9. The plot of the fraction of G+C at synonimous site in codons (GC3s) against effective codon numbers suggest no major difference in translational efficiency in the expression of glutelin multigenes.
이태호(Tae Ho Lee),김주곤(Ju Kon Kim),남백희(Baek Hie Nahm) 한국응용생명화학회 1993 Applied Biological Chemistry (Appl Biol Chem) Vol.36 No.6
To characterize the prolamines in rice cultivars, the complete coding sequences of 17 prolamine genes from the database were analyzed. According to the phylogenic analysis of the sequences, these genes could be classified into 4 groups, Group Ⅰ to Ⅳ. The multiple alignment of the deduced amino acid sequences revealed that the four groups differ from one another in chain length caused by deletion of short internal amino acids or carboxyl terminal fragments. Each group was also found to have different amino acid composition with 1, 4, 10 and 30% of sulfur containing amino acids (methionine and cysteine) in Group Ⅰ to Ⅳ prolamines, respectively. Also the isoelectric points of these groups showed the different values of 9.2, 8.2, 6.7 and 7.4. Finally, from the analysis of colon usage pattern of prolamine genes, the colon usage for arginine, serine, threonine, isoleucine, asparagine, aspartic acid, glutamic acid and cysteine were highly biased. In the analysis of the colon usage pattern, the relation of the fraction of G/C ending colons to effective colon numbers suggests the different translational efficiency in the expression of the prolamine multigenes.
흑조위축병 바이러스 RNA 를 절단하는 망치머리형 라이보자임의 제작
박종석(Jong Sug Park),김주곤(Ju Kon Kim),손성한(Seong Han Sohn),이석순(Sug Soon Lee),황영수(Young Soo Hwang) 한국응용생명화학회 1995 Applied Biological Chemistry (Appl Biol Chem) Vol.38 No.6
To develop an antiviral agent for the rice black-streaked dwarf virus (RBSDV), a hammerhead type ribozyme, which has a potential target site on the genome segment 3, was designed. Oligonucleotides for the ribozyme and its substrate were synthesized, annealed, and cloned into a plasmid pBluescript II KS(+). Ribozyme and substrate RNAs were then synthesized by in vitro transcription with T₃ RNA polymerase, obtaining RNAs in expected size, 193 and 182 nucleotides, respectively. The substrate RNA was efficiently cleaved into two fragments when incubated with the ribozyme at 55℃, while the cleavage was not detected at 37℃. In addition, the segment 3 RNA of RBSDV was also cleaved into two fragments by the same ribozyme at 55℃. Taken together, our results demonstrated that the hammerhead ribozyme has an in vitro endonucleolytic activity and may be used as an antiviral agent in transgenic plants.
박종석(Jong Sug Park),배신철(Shin Chyul Bae),김영민(Young Min Kim),백융기(Young Ki Paik),김주곤(Ju Kon Kim),황영수(Young Soo Hwang) 한국응용생명화학회 1994 Applied Biological Chemistry (Appl Biol Chem) Vol.37 No.3
Rice black-streaked dwarf virus (RBSDV), a member of the plant reoviridae fijivirus group, causes a serious damage for rice production in Korea. To characterize the RBSDV genome, virus particles were produced by feeding of planthopper (Laodelphax striatellus F.) earring RBSDV to maize plants for 2 days. In 30∼40 days after feeding, the viral particles were purified from the infected maize roots by using 10∼40% sucrose gradient centrifugation. After treatment of 10% SDS to remove the viral coat proteins, ten viral double-stranded RNAs were resolved in agrose gel electrophoresis. Total dsRNA was then used to synthesize cDNA by reverse transcriptase and a cDNA library was constructed in the λgt11 vector. The phages that contain RBSDV cDNA fragments were selected by hybridizing with the random-primed probe prepared from RBSDV dsRNAs. After subcloning of several cDNA fragments into the pUC19 plasmid mector, one clone (pRV3) was chosen for sequencing. The pRV3 clone was shown to be Located on the RBSDV genome fragment No.3 by RNA gel-blot analysis. Sequence analysis of the clone revealed that the pRV3 contains two partial open reading frames.
형질전환 토마토 식물체에서 한국형 로타바이러스 Spike 단백질 VP4의 발현
김경일,박종화,김해영,김주곤,양재명,손동화,정인식 경희대학교 생명자원과학연구원 1998 遺傳工學論文集 Vol.10 No.-
본 연구는 형질전환 토마토 식물체에서 로타바이러스의 spike 단백질인 VP4를 발현시키고자 하였다. 발아시킨 토마토 떡잎에다 pILTAB357-VP4 플라스미드를 함유하는 Agrobacterium tumefaciens를 이용하여 형질전환을 시켰다. 로타바이러스의 외피 단백질인 VP4를 coding하는 유전자는 genomic DNA PCR을 이용하여 안전하게 형질전환 토마토 줄기에 삽입되었음을 확인하였다. VP4 단백질은 형질전환 토마토 싹에서 성공적으로 발현되었으며, 그 크기는 약 88kDa으로 나타났다.