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Direct Cloning of a Xylanase Gene from Pawan-Riau Hot Spring
HAYATI Journal of Biosciences, June 2007, p 54-58
ISSN: 1978-3019
Vol. 14, No. 2
Direct Cloning of a Xylanase Gene from Pawan-Riau Hot Spring
IS HELIANTI
Center for Bioindustrial Technology, Agency for Assessment and Application of Technology,
BPPT 2 Building 15 Floor, Jalan M.H. Thamrin No 8, Jakarta 10340, Indonesia
nd th
Phone: +62-21-3169509, Fax: +62-21-3169510, E-mail: ishelianti@yahoo.co.jp
Received January 9, 2007/Accepted May 30, 2007
A functional gene containing an Open Reading Frame (ORF) encoding a β-1, 4-endoxylanase glycosyl hydrolase
family 11 was cloned directly using metagenomic PCR-cloning method from Pawan Hot Spring sample in Riau. The gene
consisted of 642 nucleotides, encoded for 213 amino acids. The amino acid sequence analysis using BLAST showed that
the gene has high homology (93%) with xylanase gene from Bacillus subtilis. The gene showed its function when it was
subcloned into an expression vector and overexpressed in E. coli. The crude extract of the recombinant enzyme had
activity for 170 U/ml at 50 C. The result of this work showed that metagenomic approach was a powerful short cut method
o
to obtain recombinant biocatalyst that was useful for industrial application.
Key words: β-1, 4-endoxylanase, metagenomic DNA, Pawan-Riau hot-spring
___________________________________________________________________________
INTRODUCTION
Indonesia), which is a potential resource of thermophilic
enzymes gene. The gene was retrieved by means of a
Polymerase Chain Reaction (PCR)-cloning, then subcloned
into pET101/D-TOPO expression vector and overexpressed
in E. coli.
β-1, 4-Endoxylanases (Xylanases) (EC .) are
enzymes that catalyze the cleavage of xylan backbone at 1-4
carbon linkage regularly to produce xylose and xylo-
oligosaccharides. Xylanase is an important enzyme for
biotechnology, because it i
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