Secretory production of bioactive recombinant human granulocyte-macrophage colony-stimulating factor by a baculovirus expression system

Lo Chun Au, Tsun Jen Liu, Horng Der Shen, Kong Bung Choo, Sheng Yuan Wang

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

The proliferation and differentiation of hematopoietic cells are stimulated by a group of glycoproteins called colony stimulating factors (CSFs). Previously, we found that the human hepatoma cell line HA22T/VGH secreted a high level of human granulocyte-macrophage colony-stimulating factor (hGM-CSF). The cDNA of hGM-CSF, including the signal peptide sequence, was amplified from the total RNA of HA22T/VGH by a reverse-transcription polymerase chain reaction and was cloned into the pUC18 vector. After confirming the nucleotide sequence, the cDNA was inserted into a pVL1393 baculovirus transfer vector. The recombinant baculovirus carrying hGM-CSF cDNA was generated by co-transfecting the hGM-CSF recombinant transfer vector and BaculoGold(TM) baculovirus DNA into the Sf9 insect cells. The expected hGM-CSF transcript was detected in the recombinant virus-infected Sf9 cells. The conditioned media of the infected cells were analyzed by a slot-blot immunoassay. The results indicate that the infected insect cells produced and secreted hGM-CSF. According to colony forming assay, a maximum titer of 2.1 x 106 U ml-1 of hGM-CSF in the medium was obtained on the third day after infection.

Original languageEnglish
Pages (from-to)107-113
Number of pages7
JournalJournal of Biotechnology
Volume51
Issue number2
DOIs
Publication statusPublished - Nov 1 1996
Externally publishedYes

Keywords

  • Baculovirus
  • Recombinant human granulocyte-macrophage colony-stimulating factor (hGM-CSF)
  • Secretion

ASJC Scopus subject areas

  • Biotechnology

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