Abstract:
A prokaryotic in situ polymerase chain reaction (PI-PCR) technique was applied to visualize Vibrio halioticoli cells using alginate lyase gene alyVG2 as a target gene. Prior to PI-PCR, a primer set, VG2-OS3, for specific amplification of an approximately 1.0-kb fragment from V. halioticoli genomic DNA was developed with amplified fragments from V. pelagius and V. fischeri DNAs as reference strains. One-stage PI-PCR using the primer set, digoxigenin-labeled dUTP, and indirect alkaline-phosphatase-linked fluorescence detection technique (HNPP/Fast Red TR as a substrate) failed to differentiate V. halioticoli IAM14596T cells from ATCC25916T cells of the closely related species V. pelagius. However, two-stage PI-PCR adding the extension and digoxigenin-labeling step of the amplified fragment into the first amplification stage allowed us to differentiate V. halioticoli cells from V. pelagius cells.
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Received May 7, 1999; accepted September 4, 1999.
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Sugimura, I., Sawabe, T. & Ezura, Y. In Situ Polymerase Chain Reaction Visualization of Vibrio halioticoli Using Alginate Lyase Gene AlyVG2 . Mar. Biotechnol. 2, 74–79 (2000). https://doi.org/10.1007/s101269900009
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DOI: https://doi.org/10.1007/s101269900009