식물홀몬을 포함한 각종 식물생장조절물질의 생검법(bioassays or biological assays)을 소개 및 비교검토함과 동시에 일관성 있는 결과를 얻기 위한 몇가지 유의점을 지적코자 옥신류 생검에서는 녹두발근과 귀리제1절간장신장, gibberellin에서는 왜성벼의 제2엽신장과 왜성완두의 상조축신장, 그리고 cytokinin류에서는 무자엽생장, 담배줄기수캘러스생장, 그리고 오이자엽내엽록소형성 등의 방법을 요약하였다. 아울러 일련의 실험을 통해 얻어졌던 각종생장조절물질(특히 최근에 개발되는 각종 왜화제)에 대한 반응을 홀몬별로 요약 및 비교하였다.
A bioassay is a test system using a living organism (in whole or in part) to determine the presence or relative potency of chemical substances. The development and uses of bioassay are intimately linked to the discovery and characterization of the major classes of plant hormones. An application of this relationship is helpful for understanding the concept of plant hormones as well as the use of bioassay. And plant bioassay have been development and employed not only for the discovery and characterization of the biological activity of plant growth regulators but also have served several important secondary roles. The ideal bioassay should possess the characteristic of high specificity. great sensitivity. short response time, low cost and ease of obtaining plant material. acceptable ease of manipulation, and minimal space and equipment requirements.
The analytic principles of GC and MS were explained in relation to plant hormone analyses and the characteristics of two instruments were compared. The selection of column, condition of measurement and the method of ionization to get a good spectrum were also briefly described. Finally, the pre-treatment of sample by solvent extraction method to remove the unnecessary part of sample and the synthetic method, especially reagents and reaction condition, for the preparation of ether or ester derivative which can be easily vaporized in GC were explained.
Ethylene gas classified as one of five major plant hormones plays an important role in various plant metabolism. The precise analysis of ethylene production of plants or plant parts is a valuable research procedure because knowledge of ethylene production facilitates measures of the physiological activity within the tissue. This paper describes procedures for analyzing ethylene from plant tissues by gas chromatography and discusses problems associated with extracting gas samples either by introducing a vacuum to plant samples or by using a hypodermic syringe. Introduced are a continuous flow system for efficient analysis and an automated system for sampling, analyzing, calculating and recording ethylene production data.
In spite of the development of highly sophisticated instrument, the precise quantitation of plant hormones still has many difficulties. Due to their high specificity, sensitivity and minimal sample purification steps, immunological assays have been widely applied for plant hormone assay. Enzme-linked immunosorbent assay technique for the determination of plant hormones was developed by Voller in 1978. Immunological assays are accomplished by competition of labeled tracer antigen and unlabeled antigen for a limited number of specific antibodies. The use of enzyme as replacement labels for radioisotopes enabled much of the sensitivity and specificity of radioimmunoassay (RIA) to be retained but without the inherent disadvantage of high capital cost, potential health hazard, and short shelf life of the labeled reactants.
식물 홀몬의 면역적 분석은 무엇보다 분석전 복잡한 정제가 필요치 않고 정밀분석을 신속히 할 수 있다는데 그 장점이 있다. 다만 pAb를 이용하는 경우에 특이성이 다소 문제로 대두되고 있으나 mAb를 생산함으로써 크게 개량할 수 있었다. 물론 GA류에 있어서는 극히 유사한 구조를 가진것 끼리의 면역분석은 잘되지 않는 것처럼 받아들여지고 있으나 이 문제도 epitope를 달리 함으로써 어느 정도 해결 팔 수 있으며, 이경우 HPLC로 정제후 mAb-ELISA를 이용하여 검출하면 훨씬 정확한 분리와 분석이 가능 할 것으로 생각된다. 본 연구자 등은 mAb를 이용한 식물 홀몬 분석에 있어서 시료를 추출, 정제후 실제 ELISA에 의해서 정량분석에 소요되는 시간은 2시간 미만정도밖에 걸리지 않는다. 또한 검출 한계도 pmol~fmol 정도로 정밀분석이 가능하다. 그리고 소요되는 장비는 간단한 spectrophometer만 가지면 된다. 다만 mAb를 생산하는 과정이 복잡하고 시간이 오래 걸린다. 따라서 필요로 하는 항체를 구입해서 (ABA artiserum sigma) 사용하는 것이 오히려 편리 할 것이다. 본 연구자 둥은 mAb를 이용한 식물 홀몬정량분석용 면역킷트를 제조하였다. 상기와 같은 여러가지 결과들은 식물 홀몬의 분석에 면역측정법이 편리하게 이용될 수 있음을 시사하고 생각된다.