Construction of a lentivector containing over-expressing β-catenin gene by multisite Gateway technology / 国际生物医学工程杂志
International Journal of Biomedical Engineering
; (6): 207-211,后插2, 2013.
Article
en Zh
| WPRIM
| ID: wpr-598760
Biblioteca responsable:
WPRO
ABSTRACT
Objective To construct a lentiviral vector over-expressing β-catenin gene by multisite Gateway technology and confirm its effect.Methods By using multisite Gateway clone technique,the entry clone of pDown-Ctnnb1 was constructed using BP recombination reaction.Then,LR recombination reaction was performed among pUp-EF1A,pDown-Ctnnb1,pTail-IRES/DsRed-Express2 and pLV.Des3d.P/puro to generate an expression clone of pLV.EX3d.P/puro-EF1A>Ctnnb1 >IRES/DsRed-Express2.In each step,PCR and sequencing analysis were used to verify the constructions.When it was verified that plasmids were transfected into 293T cells,PT-PCR was performed to determine the mRNA level of β-catenin gene.Results Both PCR and sequencing analysis revealed that β-catenin over-expression gene was inserted into the target site and the insertion sequence was perfectly corrected.The RT-PCR results showed that the expression of β-catenin gene was significantly upregulated.Conclusion The lenvivirus-mediate β-catenin over-expression gene was successfully constructed..
Texto completo:
1
Índice:
WPRIM
Idioma:
Zh
Revista:
International Journal of Biomedical Engineering
Año:
2013
Tipo del documento:
Article