Guanidinylation SS-PAAs polymer as well as preparation and application thereof
A technology of polyaminoamine and guanidinylation, which is applied in the direction of gene therapy, genetic material composition, and the use of vectors to introduce foreign genetic material, etc., to achieve the effect of improving transfection efficiency and reducing cytotoxicity
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Embodiment 1
[0063] Example 1: Preparation of SS-PAAs with Agmatine Sulfate as Guanylation Reagent
[0064] Materials and Methods
[0065] 2,2,4,6,7-Pentamethyldihydrobenzofuran-5-sulfonyl chloride (Pbf-Cl): purchased from Jill Biochemical (Shanghai) Co., Ltd. N,N-bis(acryloyl)cystamine (CBA): purchased from Alfa Aesar (Tianjin) Chemical Co., Ltd. Agmatine sulfate: purchased from Changzhou Archie Biotechnology Co., Ltd. Trifluoroacetic acid (TFA): purchased from Tianjin Kemiou Chemical Reagent Co., Ltd. pcDNA3.1-EGFP-cDNA plasmid and plasmid extraction kit: purchased from Tiangen Biochemical Technology Co., Ltd. pSliencer TM 4.1 CMV-FANCF-shRNA plasmid, FANCF-shRNA, and 328-microRNA fragment: were designed, synthesized and purchased by Shanghai Bioengineering Co., Ltd. Lipofectamine2000: purchased from Invitrogen Company; ExGen500: purchased from Fermentas Company; MTT assay cell viability kit, cell experiment consumables, culture medium, agarose, etc.: all purchased from Baoxin Biolo...
Embodiment 2
[0073] Implementation example 2: prepare SS-PAAs with chlorhexidine acetate monomer as guanidinylation reagent
[0074] (1) Protection of Pbf-Cl on the guanidinium structure in chlorhexidine acetate
[0075] Put 1g of chlorhexidine acetate in a 50ml three-neck bottle, add 15ml of water to dissolve, cool down to below 0°C, and adjust the pH to 11; slowly add 2g of Pbf-Cl dissolved in 10ml of acetone dropwise, and the temperature is maintained at 0°C Below, the pH is maintained in the range of 10-13; after the dropwise addition, react below 0°C for 1 hour and at room temperature for 3 hours until the pH does not change. The progress of the reaction was monitored by TLC. Suction filtration, rotary evaporation to remove acetone in the filtrate, collect the acetone phase product, oven dry at 40-60°C overnight, weigh, and obtain chlorhexidine whose guanidine structure is protected by Pbf-Cl.
[0076] (2) Synthesis of guanidinated SS-PAAs polymer with chlorhexidine as monomer
[0...
Embodiment 3
[0081] Implementation example 3: Preparation of SS-PAAs with guanidine hydrochloride monomer as guanidinylation reagent
[0082] (1) Protection of Pbf-Cl on the guanidinium structure in guanidine hydrochloride
[0083] Put 1g of guanidine hydrochloride in a 100ml three-neck flask, add 5ml of water to dissolve, cool down to below 0°C, adjust the pH to 11; slowly add 3.02g of Pbf-Cl dissolved in 10ml of acetone dropwise, and maintain the temperature at 0°C Below, the pH is maintained in the range of 10-13; after the dropwise addition, react below 0°C for 1 hour and at room temperature for 1 hour until the pH does not change. The progress of the reaction was monitored by TLC. Suction filtration, collect the precipitated product, dry in an oven at 40-60°C overnight, and weigh to obtain guanidine hydrochloride whose guanidine structure is protected by Pbf-Cl.
[0084] (2) Synthesis of guanidinated SS-PAAs polymer with guanidine hydrochloride as monomer
[0085] Take 0.19g of the...
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