Application of chylomicron as liver targeting gene treatment vector
A chylomicron, gene therapy technology, applied in gene therapy, non-effective components of polymer compounds, genetic material components, etc., can solve the problems of large cytotoxicity and weak carrier specificity, etc.
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Embodiment 1
[0023] Example 1: Preparation of a dual-targeting gene vector in which chylomicrons encapsulate the pAFP-TK-IRES2-EGFP plasmid vector.
[0024] Prepare the pAFP-TK-IRES2-EGFP plasmid vector with a concentration of 50 μg / mL, place the chylomicron in a sonicator and sonicate 10 times, each time for 15 s, with an ultrasonic power of 150 W, according to the mass volume ratio of the plasmid vector to chylomicron 2 μg / mL The plasmid vector and chylomicron were mixed, vortexed for 30 seconds, and left to stand for 30 minutes to prepare a dual-targeting gene carrier in which the chylomicron encapsulated the pAFP-TK-IRES2-EGFP plasmid vector.
[0025] The eukaryotic expression plasmid vector pAFP-TK-IRES2-EGFP was purchased from Beijing Xiercheng Industrial Technology Co., Ltd., and the plasmid map of the plasmid vector is as follows: figure 1 , which contains the alpha-fetoprotein (AFP) promoter for gene expression, the AFP promoter can only be activated under a specific AFP activatio...
Embodiment 2
[0026] Example 2: Encapsulation effect of chylomicron on pAFP-TK-IRES2-EGFP plasmid vector.
[0027] The dual-targeting carrier obtained in Example 1 was observed by scanning electron microscopy, and the 100,000-fold scanning electron microscope image showed that chylomicrons had a good encapsulation effect on the pAFP-TK-IRES2-EGFP plasmid vector, and the particle size was between 200 and 300 nm. ,Such as figure 2 .
Embodiment 3
[0028] Example 3: Protective effect of chylomicron on pAFP-TK-IRES2-EGFP plasmid vector.
[0029] Bovine pancreatic deoxyribonuclease I was added to the dual-targeting vector obtained in Example 1 to make the final concentration 20 μg / mL, and digested at 37°C for 30 minutes. The simple pAFP-TK-IRES2-EGFP plasmid vector was used as a control. The protective effect of chylomicron on pAFP-TK-IRES2-EGFP plasmid vector was observed by agarose gel electrophoresis. Observation see image 3 , Lane 1 shows that the simple pAFP-TK-IRES2-EGFP plasmid vector is degraded by bovine pancreatic deoxyribonuclease Ⅰ because there is no protective effect of chylomicrons; Lane 2 shows the effect of chylomicrons on pAFP-TK-IRES2 The -EGFP plasmid vector has a strong protective effect and can avoid its degradation by bovine pancreatic deoxyribonuclease Ⅰ. The results showed that the chylomicrons in the dual-targeting vector had a strong protective effect on the pAFP-TK-IRES2-EGFP plasmid vector a...
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