Invigentech高效率轉(zhuǎn)染DNA siRNA到免疫T細(xì)胞
一、浙江大學(xué)附屬第二醫(yī)院乳腺外科;浙江大學(xué)附屬第二醫(yī)院浙江省療法腫瘤微環(huán)境與免疫重點(diǎn)實(shí)驗(yàn)室;浙江省人民醫(yī)院,浙江省人民醫(yī)院,杭州醫(yī)學(xué)院附屬人民醫(yī)院,浙江省個(gè)體化醫(yī)學(xué)腫瘤分子診斷與診斷重點(diǎn)實(shí)驗(yàn)室;紹興大學(xué)附屬醫(yī)院甲狀腺乳腺外科在2020-2-19聯(lián)合發(fā)表標(biāo)題為Breast cancer-derived exosomes transmit lncRNA SNHG16 to induce CD73+γδ1 Treg cells(乳腺癌衍生的外泌體傳遞lncRNA SNHG16誘導(dǎo)CD73 +γδ1 Treg細(xì)胞)的文章到nature/sigtrans,文章已被接受;
二、本研究使用中使用Invigentech(英克)公司INVI DNA RNA 轉(zhuǎn)染試劑轉(zhuǎn)染質(zhì)粒DNA、siRNA到Vδ1 T 免疫T細(xì)胞里面;
三、本文中轉(zhuǎn)染質(zhì)粒DNA、siRNA,轉(zhuǎn)染細(xì)胞數(shù)量信息如下:
A. 將全長2435 bp的序列克隆到pCR3.1載體中構(gòu)建SNHG16過表達(dá)載體;
B. siRNA序列:SNHG16-homo-349,5′GCCUCUGCUGCUAAUUGUUTT-3'; SNHG16-homo-867, 5′-CCAAGGAGGGACUGUUUAATT-3′和SNHG16-homo-2004,5′-CCCAGUGUUGACUCACCAATT-3;
C. 轉(zhuǎn)染細(xì)胞用量:6孔板里面1 × 106 cells/well;
四、發(fā)表文章部分內(nèi)容如下:
Breast cancer-derived exosomes transmit lncRNA SNHG16 to induce CD73+γδ1 Treg cells
Plasmid construction and siRNA silencing A full-length 2435 bp sequence was cloned into the pCR3.1 vector to
construct the SNHG16 overexpression vector. The sequences of SNHG16-specifific siRNAs were as follows: SNHG16-homo-349, 5′GCCUCUGCUGCUAAUUGUUTT-3′; SNHG16-homo-867, 5′-CCAAGGAGGGACUGUUUAATT-3′ and SNHG16-homo-2004, 5′-CCCAGUGUUGACUCACCAATT-3′. The miR-16–5p mimics, inhibitor and negative
controls were purchased from GenePharma (SupplementaryTable S3).
To manipulate the expression of miR-16–5p in Vδ1 T cells, Vδ1T cells were sorted from peripheral blood and seeded into six-wellplates at 1 × 106 cells/well with 1 ml of medium supplementedwith 10% FBS, 10 μg/ml CD3, 10 μg/ml CD28 and 40 U/mL IL-2.Then, transfection reagent (INVI DNA RNA Transfection Reagent,Invigentech, USA) and miR-16–5p mimics/NC/inhibitor/inhibitor NC (20 μm) were incubated with the cells for 48 h, after which the cells were collected for further experiments.
but not Vδ2+ T cells were revealed to comprise the majority of TILs in all three BC molecular subtypes when gated on CD3 (p < 0.01, Fig. 1b, c). In addition, immunoflfluorescence of frozen sections further confifirmed the dominant distribution of Vδ1+T cells in BC (Fig. 1d).
To further identify the function of breast cancer-infifiltrating γδ1T cells, BC cell lines (MCF-7, T-47D, MDA-MB-231 and MDA-MB-468) were co-cultured with freshly isolated Vδ1+ T cells frombreast tumour tissue (E:T ratios: 1:1, 5:1 and 10:1). Upon plotting the cell growth curve, the data show that γδ1 T cells did not exert
invigentech(英克)INVI DNA RNA轉(zhuǎn)染試劑信息如下
產(chǎn)品名稱 | 貨號 | 規(guī)格 | 報(bào)價(jià) |
INVI DNA RNA 轉(zhuǎn)染試劑 | IV1216025 | 0.25 ml | 詢價(jià) |
INVI DNA RNA 轉(zhuǎn)染試劑 | IV1216050 | 0.50 ml | 詢價(jià) |
INVI DNA RNA 轉(zhuǎn)染試劑 | IV1216075 | 0.75 ml | 詢價(jià) |
INVI DNA RNA 轉(zhuǎn)染試劑 | IV1216100 | 1.00 ml | 詢價(jià) |
INVI DNA RNA 轉(zhuǎn)染試劑 | IV1216150 | 1.50 ml | 詢價(jià) |
INVI DNA RNA 轉(zhuǎn)染試劑 | IV1216300 | 3.00 ml | 詢價(jià) |
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