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KMID : 0613820040140050840
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Journal of Life Science
2004 Volume.14 No. 5 p.840 ~ p.846
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Expression of Recombinant Korean Mistletoe(KM) Lectin and B genes in Saccharomyces cerevisiae
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Choi Yoon-Hyeok
Kim Jong-Bae
Yang Woong-Suk
Hwang Cher-Won
Shi Wen-Tao
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Abstract
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A study for expression of Korean Mistletoe (KM) lectin gene (A,B) in Saccharomyces cerevisiae was done using transforming system of yeast. In order to overexpress the genes efficiently in yeast, two lectin genes (A,B) were re-cloned and modified including Kozak translation initiation sequence using PCR amplification. The constructed plasmids containing modified lectin A and B genes were transformed to S. cerevisea INVSc (MATα, his3△1, leu2, trpl-289, ura3-52). The transformed cells were identified by DNA sequencing with ABI3700 system and induced with 2% of galactose for recombinant KM lectin (rKM lectin) protein. The rKM lectin A and B proteins were determinated about 29kDa size of protein by SDS-PAGE and western blotting analysis. The expressed recombinant lectin was determinated 1.24～1.75 μg per 1 mg of cytosolic soluble protein by sandwich ELISA method. Moreover the lectin genes were expressed as maximum level at 36 h after galactose induction and lectin A gene was were repressed after 48 h.
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KEYWORD
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Korean Mistletoe(KM), recombinant lectin, recombinant yeast, RIP
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