peg-acs/m-siRNA nanocomposite and its application and method for reducing histamine content in fishmeal storage
A PEG-ACS, nanocomposite technology, applied in the field of feed industry, can solve the problems of high cytotoxicity and in vivo toxicity, low efficiency, weak targeting, etc. Effect
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Embodiment 1
[0057] (1) Design the M-siRNA sequence.
[0058] Sense strand: 5'-GGAUCGUAUUUCUGUUGAAAU-3',
[0059] Antisense strand: 5'-UUCAACAGAAAUACGAUCCAC-3'.
[0060] (2) Preparation of M-siRNA.
[0061] (3) Preparation of ACS: Weigh 0.5g chitosan, dissolve it in TEMED / HCl buffer solution with pH 5.0, then add 567mg EDC and 340mg NHS coupling agent to the solution, stir well, then add Arginine with 80% molar weight of the polysaccharide amino group was reacted at room temperature for 8 hours under magnetic stirring, and then desalted water dialysis and freeze-drying were performed to obtain arginine-modified chitosan.
[0062] (4) Preparation of PEG-ACS: Take 2.5 mL of 10 mg / mL arginine-modified chitosan, add 25 mg of PEG-SPA, and react at room temperature for 4 h. Use a dialysis bag with a molecular weight cut-off of 14,000 to remove unreacted PEG. The obtained PEGylated arginine-modified chitosan was freeze-dried for future use.
[0063] (5) Preparation of siRNA solution: siRNA w...
Embodiment 2
[0075] (1) Design the M-siRNA sequence.
[0076] Sense strand: 5'-GGAUCGUAUUUCUGUUGAAAU-3',
[0077] Antisense strand: 5'-UUCAACAGAAAUACGAUCCAC-3'.
[0078] (2) Preparation of M-siRNA.
[0079] (3) Preparation of ACS: Weigh 0.8g chitosan, dissolve it in TEMED / HCl buffer solution with pH 4.5, then add 900mg EDC and 550mg NHS coupling agent to the solution, stir well, then add Arginine with an equimolar amount of polysaccharide amino groups was reacted at room temperature for 7 hours under magnetic stirring, and then desalted water dialysis and freeze-drying were performed to obtain arginine-modified chitosan.
[0080] (4) Preparation of PEG-ACS: Take 3 mL of 8 mg / mL arginine-modified chitosan, add 24 mg of PEG-SPA, and react at room temperature for 4 h. Use a dialysis bag with a molecular weight cut-off of 14,000 to remove unreacted PEG. The obtained PEGylated arginine-modified chitosan was freeze-dried for future use.
[0081] (5) Preparation of siRNA solution: siRNA was ...
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