[Construction and expression of a fusion protein containing extracellular domain of human Delta-like1 and Fc fragment of human IgG1 in the Pichia pastoris].
OBJECTIVE
To construct a Pichia pastoris expression vector PIC- hDll1(ext)-Fc, and to express the fusion protein containing the extracellular domain of human Delta-like1 and the Fc fragment of human IgG1 fusion gene in Pichia pastoris.


METHODS
The extracellular domain of human Delta-like1 gene was amplified from pEF-BOSneo-hdll1(ext)-Fc by PCR. The expression vector was constructed by DNA recombination. The constructed plasmid was transformed into yeast GS115 by electroporation. The recombinant transformants with a high copy number of the plasmid were selected by using MD plate and G418. The expression of protein was induced by addition of methanol.Then analyzed protein expression by SDS-PAGE and Western blot.


RESULTS
The extracellular domain of human Delta-like1 gene was effectively amplified. The DNA sequencing result showed that the constructed plasmid containing hDll(ext)-Fc fusion gene was the same as designed. The fusion protein was successfully expressed in Pichia pastoris.


CONCLUSIONS
The hDll1(ext) gene has been successfully cloned and expressed in the form of Fc fusion protein, which provides a new fusion protein for further experiments.
Huang
Si-yong
SY
Department of Hematology, Tangdu Hospital, Fourth Military Medical University, Xi'an 710038, China. huangsiyong1980@126.com
Wu
Yan-lan
YL
Li
Guo-hui
GH
He
Fei
F
Kang
Zhi-jie
ZJ
Liu
Li
L
Han
Hua
H
Liang
Ying-min
YM
D018274:Electroporation; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000235:genetics / Q000378:metabolism*; D007074:Immunoglobulin G / Q000235:genetics / Q000378:metabolism; D000432:Methanol / Q000737:chemistry; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins / Q000494:pharmacology*
D005656:Fungal Proteins; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D000432:Methanol