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題 名 | 以聚合酶鏈反應--限制酶水解反應鑑定Rhodococcus屬菌株=Identification of Rhodococcus Species by Polymerase Chain Reaction-Restriction Enzyme Digestion |
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作 者 | 程梅萍; 廖仁寶; 王西華; | 書刊名 | 中國農業化學會誌 |
卷 期 | 37:3 1999.09[民88.09] |
頁 次 | 頁328-337 |
分類號 | 369.4 |
關鍵詞 | 鑑定; 聚合酶鏈反應; 限制酶水解; Rhodococcus; Nocardia; Polymerase chain reaction; Restriction enzyme digestion; |
語 文 | 中文(Chinese) |
中文摘要 | Rhodococcus屬菌株能行廣泛的代謝途徑,常被應用於分解污染物。但是,此屬菌株傳統鑑定法相當困難且費時。本研究目的即為發展聚合□鏈反應法,以快速鑑定 Rhodococcus屬菌株。本研究由Rhodococcus spp. 16S rRNA基因的變異區設計引子rh16s、rh16s5及rh16s6,以專一性增幅Rhodococcus spp. 16S rRNA基因部分序列,來鑑定Rhodococcus spp.。使用此引子對進行聚合□鏈反應後,Rhodococcus spp.及類綠關係相近的Nocardia asteriodes及N. carnea,皆得到大小如預期的820 bp之聚合□鏈反應產物。但是,其餘16株格蘭氏陰性及5株格蘭氏陽性土壤常見對照菌株則無聚合□鏈反應產物產生。為區別Rhodococcus 與Nocardia屬,將兩者的聚合□鏈反應產物與限制□Pvul反應,結果顯 示N. asteriodes及N. carnea的聚合□鏈反應產物會被水解為兩片段,其大小各約380、440by;Rhodococcus屬則否。因此,聚合□鏈反應及其後的限制□Pvul水解反應可以用於快速鑑別Rhodococcus spp.。應用此鑑定法,菌株不須經培養,較傳統方法快速、簡便且靈敏度高。 |
英文摘要 | The genus Rhodococcus is a unique taxon consisting of microorganisms that exhibit a broad metabolic pathway, which can be applied to degrade various pollutants. The identification of Rhodococcus species is difficult and time-consuming by the conventional method. In this study, the PCR primers rh16s, rh16s5 and rh16s6 for the amplification of partial 16/s rRNA genes of Rhodococcus spp. were designed according to the sequence of the variable region of 16S rRNA genes from Rhodococcus spp. With these primers, DNA fragments with predicted sizes were amplified from Rhodococcus spp. and phylogenetically-related Nocardia asteriodes and N. carnea, but not from 16 other strains of Gram-negative or strains of Gram-positive microorganisms. In order to differentiated Rhodococcus spp. from Nocardia spp., the PCR products of both genera were reacted with a restriction enzyme, Pvul. It was found that the PCR products of Nocardia asteriodes and N. carnea were digested into two fragments while those of Rhodococcus spp. were not. Thus, the genus Rhodococcus could be quickly identified by means of PCR and subsequent restriction enzyme digestion. |
本系統中英文摘要資訊取自各篇刊載內容。