Embedded tissue-specific expressed Cre tool mice built by utilizing Flp-Frt system
A flp-frt, cd4-stop-cre technology, applied in the biological field, can solve the problems such as not many Cre transgenic tool mice, incomplete tissue-specific promoters, affecting expression, etc.
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[0044] One, the construction of the tool vector pBS-Pr-Flp of eukaryotic expression Flp recombinase (flow process sees Figure 4 ), including the following steps:
[0045] Step 1, with Flp-ER (129S4 / SvJaeSor-Gt (ROSA) 26Sor tm1(FLP1)Dym / J, stock No.003946, Jackson Laboratory) mouse genome DNA was used as a template, and the Flp fragment was amplified by PCR. Primers are: FlpF:CG GGATCCGCCGCC ATGGGGATGCCACAATTTGATATATTATG; FlpR:CG ACGCGT TTATATGCGTCTATTTATGTAGG. The underlined part is the restriction site introduced by PCR. The PCR product was connected to pMD18-T and identified by enzyme digestion. The clone with the correct enzyme digestion identification result was sent to Jinsite Technology (Nanjing) Co., Ltd. for sequencing to confirm the correctness. After the Flp fragment was ligated from pMD18T with restriction endonucleases BamHI and MluI, the Flp fragment was substituted for the Cre fragment in the pBS185 plasmid (Gibico), and the digestion was verified to be...
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