Construction of Novel Phytochelatins by Overlap Oligonucleotides
Keywords:Design of oligonucleotide, Hexahistidine, Synthetic phytochelatin
Synthetic phytochelatins are protein analogs of phytochelatin with similar heavy metal binding affinities that can be easily produced from a synthetic DNA template. We design synthetic phytochelatin [(Glu-Cys)n Gly] linked to hexahistidine by viral linker peptide and then followed by gene synthesis and cloning of it. Then peptide coding gene (synthetic phytochelatin with linker and hexahistidine) was designed exactly and constructed with step by step methods by overlapping oligonucleotides using T4 DNA Ligase. Finally, synthesized gene amplified by PCR, cloned in pTZ57R/T and transformed to Escherichia coli (DH5α). The results of sequencing show that some types of synthetic phytochelatin (EC4, EC12, and EC20) with linker and hexahistidine were constructed and cloned in vector.
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