| Summary: | 碩士 === 中山醫學大學 === 生物化學研究所 === 90 === Abstract
First we adopted Gram stain method to examine the fermented milk. As the result we found a total of 3 bacteria, namely Gram’s positive bacilli, positive coccus, and yeast. In order to cultivate BAP medium and EMB medium were utilized and the three bacteria were cultured under aerobic eniverment. Incidentally the specimen were also cultured in no anaerobic environment condition of BAP, KVLB, BBE media but no bacteria was found under this condition. In BAP medium there are 3 kinds of bacteria have been found to grow. Again gram stain proved they are gram’s positive coccus, α-hemolytic Gram’s positive bacilli and yeast: also in EMB medium Gram’s positive bacilli and yeast were found to grow. Followed use Biolog System inspection they are respectively identified as Leuconostoc citrium and cryotcoccus albidus var aerius.. However,α-hemolytic Gram’s positive bacilli failed to be identified by the Biolog System. This shows that the Biolog System may not be able to identify all bacteria of this kind. Thus, the conventional way of using Biochemical tests identified α-hemolytic Gram’s positive bacilli is Lactobacilus Sp.
In the past the assessment of relationship between two species of bacteria was done by random expansion DNA section multi status analysis which is the most quick, easy and effective assessment method available. This analysis method has been used since 1990, and widely used in differentiation of DNA of various species of bacteria., and also used in identifying relationship between fungus species. Utilizing PCR-RFLP, RAPD-PCR and portion of 16S rRNA to make sequence analysis has helped to understand the difference between bacteria to proceed in identify bacteria species, to establish a DNA bank. Furthermore, based on utilizing 16S rRNA sequential analysis not only identify known bacteria(4,38)but also unknown bcteria and its position (28,34). Therefor, in this study applied PCR and 16S rRNA to different species and made sequential analysis. In recent years using 16S rRNA gene sequence a lot of new subspecies and species were located. In this study thus we use this technique to identify bacteria and fungus and in order to effectively extract RNA, finally a medium offered by Biolog
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System was used. In 16S rRNA gene highly conserved region preservation section a pair of primer (44) was installed. Then using PCR method and amplify 16S rRNA gene, and again use the primer to proceed in gene sequential analysis. The result of the analysis outcome was compared with that of gene bank it was found that there was more than 98% similarity exist in DNA sequences between the separated Lactobacillus sp. and Leuconostoc citrium sp. versus those of known bacteria.It is thus confirmed that separated bacteria belonged to Lactobacillus delbtueckii subsp. Lactis, Leuconostoc citrium. As to crypotococcus albidus var aerius, up to date, obviously not used in fermentation of dairy products. As far as we know that cryptococcus albidus var aerius existed in skin, soil and birds. Therefore it is considered to be a pollution organism or may pollute dairy products during the production processes.
Many dairy products in market such as yogurt, yogurt milk, yangloto etc.often claim that they can enhance human health, and moreover wanted to improve it by genetic engineering that transplant special gene to increase utilization value of lactic acid bacilli.
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