Isolation of polyhydroxyalkanoates-producing bacteria using a combination of phenotypic and genotypic approach.
OBJECTIVE
To develop an efficient approach using a combination of phenotypic and genotypic methods for isolation of environmental bacteria that produce mid-chain-length polyhydroxyalkanoates (mcl-PHAs).


RESULTS
A viable-colony staining method using Nile red was used to screen for PHA-producing bacteria followed by a polymerase chain reaction (PCR) screen using primers to amplify the partial nucleic acid sequence of the phaC1 synthase gene for confirmation. Microbes containing lipophilic storage compounds isolated from environmental samples could readily be detected by the colony staining method. They were further examined by Sudan Black staining to highlight the inclusions inside the cells. These isolates were subsequently subjected to PCR analysis. As a result, more than a hundred strains were identified as PHA-positive isolates from this screening approach.


CONCLUSIONS
These results conclusively demonstrate that environmental bacterial strains able to accumulate the PHAs could readily be obtained by this screening method.


CONCLUSIONS
We propose a polyphasic approach using a combination of phenotypic and genotypic screening method to rapidly screen and identify bacteria able to produce significant amounts of mcl-PHAs from environment. This approach can be adopted as a rapid screen for micro-organisms able to accumulate PHAs to be used for potential manufacture and other industrial applications.
Kung
S-S
SS
Graduate School of Biotechnology and Bioinformatics, Yuan Ze University, Chung-Li, Taiwan.
Chuang
Y-C
YC
Chen
C-H
CH
Chien
C-C
CC
D000217:Acyltransferases / Q000737:chemistry / Q000235:genetics; D000595:Amino Acid Sequence; D001419:Bacteria / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D005798:Genes, Bacterial; D005838:Genotype; D007220:Industrial Waste; D008969:Molecular Sequence Data; D010641:Phenotype; D010802:Phylogeny; D011091:Polyesters / Q000302:isolation & purification / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D012988:Soil Microbiology
D004269:DNA, Bacterial; D007220:Industrial Waste; D011091:Polyesters; D000217:Acyltransferases; C077225:poly(3-hydroxyalkanoic acid) synthase