Synthesis of a wild-type and three mutant Cucurbita maxima trypsin inhibitor-encoding genes by a single-strand approach.
A single-strand approach to gene assembly, based on a modification of an in vitro complementary oligodeoxyribonucleotide template-directed ligation of the desired sequence to a linearized vector [Chen et al., Nucleic Acids Res. 18 (1990) 871-878], is described. The gene coding for the wild-type Cucurbita maxima trypsin inhibitor of 29 amino acid residues [Bode et al., FEBS Lett. 242 (1989) 285-292], as well as three mutant forms of the gene, in which two of the three disulfide bonds have been replaced singly or as a pair, have been synthesized in a single synthesis run with minimal manual intervention. Subsequent to ligation to pUC9 and in vivo gapped duplex repair by Escherichia coli, their sequences have been verified.
Department of Biochemistry, University of Cape Town, Rondebosch, South Africa.
D P
DP
Botes
M D
MD
Qobose
V A
VA
Corfield
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004277:DNA, Single-Stranded; D008969:Molecular Sequence Data; D009154:Mutation; D010944:Plants / Q000235:genetics; D010957:Plasmids; D014361:Trypsin Inhibitors / Q000096:biosynthesis* / Q000235:genetics
D004277:DNA, Single-Stranded; D014361:Trypsin Inhibitors