[The construction and identification of the PRNP vectors with ubiquitin or the lysosome-targeting signal].
OBJECTIVE
To evaluate PrP expression characteristic of PRNP nucleic acid vaccine vector with ubiquitin or the lysosome-targeting signal.


METHODS
The gene of ubiquitin and lysosome-targeting signal were ligated to PRNP and pcDNA3.1 vector that is, pcDNA3.1-UPrP and pcDNA3.1-PrPL were constructed. The expression characteristics of PrP with two signals were evaluated by Western Blot and the localization was observed by indirect immune fluorescence.


RESULTS
The protein expressed by pcDNA3.1-UPrP and pcDNA3.1-PrPL with ubiquitin and lysosome-targeting signal can be recognized by prion-specific antibody. The protein has three glycosylation molecules form as native PrP.PrP with ubiquitin was degraded gradually with time extension,whereas quantity of PrP with lysosome signal reduced in 48 h after transfection. The protein with two location signals can direct fusion proteins to cytoplasm.


CONCLUSIONS
The PRNP vectors with ubiquitin or the lysosome-targeting signal were constructed and expressed in eukaryocyte successfully. There will be one of good foundation on PRNP nucleic acid vaccine.
Li
Yuan
Y
Medical School, Anhui University of Science & Technology, Huainan 232001, China.
Mei
Guo-yong
GY
Jiang
Hui-ying
HY
Wang
Gui-rong
GR
Tian
Chan
C
Chen
Cao
C
Wang
Xin
X
Wang
Ke-xia
KX
Han
Jun
J
Dong
Xiao-ping
XP
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D019556:COS Cells; D002522:Chlorocebus aethiops; D006224:Cricetinae; D003412:Cricetulus; D005822:Genetic Vectors / Q000235:genetics / Q000276:immunology*; D006801:Humans; D008247:Lysosomes / Q000737:chemistry*; D000072002:Prion Proteins; D011328:Prions / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology*; D014162:Transfection; D025801:Ubiquitin / Q000235:genetics / Q000276:immunology*
C512656:PRNP protein, human; D000072002:Prion Proteins; D011328:Prions; D011994:Recombinant Proteins; D025801:Ubiquitin