| Summary: | Soft rot is one of the most important diseases of orchids caused by
bacterial pathogens. Bacterial soft rots can be identified by alot of methods. One
of the most prominent bacterial pathogens identification methods is nucleotide
analysis based on 16S rRNA gene sequence. Optimization of PCR technique to
amplify 16S rRNA gene is essential for time and material effieciency, that will
make identification to be rapid and more appropriate. The purposes of this study
were to (1) decide concentration of DNA and primer, and also concentration of
bacterial pure cultures and primer to amplify clear 16S rRNA gene fragment and
(2) identify bacterial soft rot of orchid based on 16S rRNA gene sequence.
Optimization of PCR was done by using variations of DNA, pure cultures of
bacteria, and primer to amplify the 16S rRNA gene sequence. Identification of
orchids bacterial soft rot was conducted using nucleotide analysis by BLAST
programme at web NCBI to know similarities coevicient between isolates and
reference strains in gene bank. The result showed that the most optimal
concentration to amplify 16S rRNA gene sequence at DNA and primer
concentration were 63,4 ng/μl and 10 pmol, also pure cultures and primer
concentration were 8 x 109 CFU/ml dan 10 pmol. Based on BLAST analysis at the
gene bank, the bacterial soft rot of orchids have similarities 99-100% with several
genera of bacteria such as Citrobacter, Enterobacter, Klebsiella, Pectobacterium,
Providencia, and Pseudomonas.
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