Amplification and Direct Sequence Analysis of the 23S rRNA Gene from Thermophilic Bacteria | Zendy

Ashraf S. Ibrahim | Zendy; Jacob Hofman-Bang | Zendy; Birgitte K. Ahring | Zendy

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Amplification and Direct Sequence Analysis of the 23S rRNA Gene from Thermophilic Bacteria

Author(s) -

Ashraf S. Ibrahim,

Jacob Hofman-Bang,

Birgitte K. Ahring

Publication year - 2001

Publication title -

biotechniques

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 0.617

H-Index - 131

eISSN - 1940-9818

pISSN - 0736-6205

DOI - 10.2144/01302rr05

Subject(s) - bacteria , biology , thermophile , dna extraction , exonuclease , alkaline lysis , gene , dna , sequence analysis , lysis , microbiology and biotechnology , polymerase chain reaction , genetics , plasmid , polymerase , dna vaccination

We present a simplified and fast method to obtain high-quality sequences directly from PCRs without the traditional gel purification. We also report on an improved method to obtain sequence-quality PCR products from microorganisms that are difficult to lyse with no need for DNA extraction. The technique uses exonuclease 1 and shrimp alkaline phosphatase to degrade residual dNTPs and primers. Our technique is shown to work on both Gram-positive and Gram-negative bacteria.

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