CLONING, CHARACTERIZATION, AND EXPRESSION OF 2 ALPHA-AMYLASE GENES FROM ASPERGILLUS-NIGER VAR AWAMORI

被引:50
作者
KORMAN, DR
BAYLISS, FT
BARNETT, CC
CARMONA, CL
KODAMA, KH
ROYER, TJ
THOMPSON, SA
WARD, M
WILSON, LJ
BERKA, RM
机构
[1] GENENCOR INC, 180 KIMBALL WAY, San Francisco, CA 94080 USA
[2] SAN FRANCISCO STATE UNIV, SAN FRANCISCO, CA 94132 USA
关键词
α-amylase; Filamentous fungi; Gene duplication;
D O I
10.1007/BF00312611
中图分类号
Q3 [遗传学];
学科分类号
071007 ; 090102 ;
摘要
Using synthetic oligonucleotide probes, we cloned genomic DNA sequences encoding an α-amylase gene from Aspergillus niger var. awamori (A. awamori) on a 5.8 kb EcoRI fragment. Hybridization experiments, using a portion of this cloned fragment to probe DNA from A. awamori, suggested the presence of two α-amylase gene copies which were subsequently cloned as 7 kb (designated as amyA) and 4 kb (amyB) HindIII fragments. DNA sequence analysis of the amyA and amyB genes revealed the following: (1) Both genes are arranged as nine exons and eight introns; (2) The nucleotide sequences of amyA and amyB are identical throughout all but the last few nucleotides of their respective coding regions; (3) The amyA and amyB genes from A. awamori share extensive homology (≥98% identity) with the genes encoding Taka-amylase from A. oryzae. In order to test whether both amyA and amyB were functional in the genome, we constructed vectors containing gene fusions of either amyA and amyB to bovine prochymosin cDNA and used these vectors to transform A. awamori. Transformants which contained either the amyA- or amyB-prochymosin gene fusions produced extracellular chymosin, suggesting that both genes are functional. © 1990 Springer-Verlag.
引用
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页码:203 / 212
页数:10
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