本文關(guān)鍵詞：脂質(zhì)體介導(dǎo)轉(zhuǎn)染的綠色熒光蛋白基因在骨髓基質(zhì)細(xì)胞內(nèi)表達(dá)　出處：《山東大學(xué)》2005年碩士論文　論文類(lèi)型：學(xué)位論文

  更多相關(guān)文章： 細(xì)胞培養(yǎng) 骨髓基質(zhì)細(xì)胞 綠色熒光蛋白 基因轉(zhuǎn)染 脂質(zhì)體

【摘要】：[目的]骨髓基質(zhì)細(xì)胞(bone marrow stromal cells,BMSC),也被稱(chēng)為骨髓間充質(zhì)干細(xì)胞(bone marrow mesenchymal cells),是存在于骨髓基質(zhì)中一種成體干細(xì)胞,來(lái)源于中胚層,具有很強(qiáng)的自我復(fù)制和多向分化功能,可以分化為多種不同的組織細(xì)胞,是細(xì)胞工程學(xué)和組織工程學(xué)研究中的一種理想種子細(xì)胞。如果BMSC容易接受外源性基因而且穩(wěn)定表達(dá),結(jié)合其多向分化能力,BMSC將會(huì)在組織工程、細(xì)胞移植、基因治療等領(lǐng)域得到十分廣闊的應(yīng)用。本實(shí)驗(yàn)的主要目的為: 1.建立一種體外分離、培養(yǎng)擴(kuò)增大鼠骨髓基質(zhì)細(xì)胞的方法,為進(jìn)一步研究和利用該細(xì)胞奠定基礎(chǔ); 2.研究脂質(zhì)體轉(zhuǎn)染法對(duì)骨髓基質(zhì)細(xì)胞進(jìn)行基因修飾的可行性和有效性,為研究骨髓基質(zhì)細(xì)胞在組織工程和基因治療中的應(yīng)用提供。 [方法]用淋巴細(xì)胞分離液分離出大鼠骨髓基質(zhì)細(xì)胞,并用含20%(體積分?jǐn)?shù))胎牛血清的DMEM培養(yǎng)液培養(yǎng),胰酶消化傳代,擴(kuò)增大鼠骨髓基質(zhì)細(xì)胞。取第3～5代BMSC,用脂質(zhì)體轉(zhuǎn)染法將外源基因綠色熒光蛋白基因介導(dǎo)進(jìn)入到骨髓基質(zhì)細(xì)胞內(nèi),然后在熒光顯微鏡下觀察,計(jì)數(shù)表達(dá)熒光蛋白細(xì)胞的數(shù)量,并用臺(tái)盼蘭排斥試驗(yàn)檢測(cè)轉(zhuǎn)染細(xì)胞的活力。 [結(jié)果] 1 大鼠BMSC的分離培養(yǎng) 用Percoll淋巴細(xì)胞分離液對(duì)大鼠骨髓作密度梯度離心,所得細(xì)胞為比較一致的球形單個(gè)核細(xì)胞,臺(tái)盼蘭染色細(xì)胞活力達(dá)99%。接種24小時(shí)后,倒置顯微
[Abstract]:[Objective to study the bone marrow stromal cells (BMSCs) of bone marrow stromal cells (BMSCs). Bone marrow mesenchymal cells, also known as bone marrow mesenchymal stem cells, is a kind of adult stem cells existing in bone marrow stromal cells, derived from mesoderm. It has a strong function of self-replication and multi-differentiation, and can differentiate into a variety of different tissue cells. It is an ideal seed cell in cell engineering and tissue engineering. If BMSC is easy to accept exogenous gene and express stably, it will be in tissue engineering if combined with its multidirectional differentiation ability. Cell transplantation, gene therapy and other fields have been widely used. 1. To establish a method of isolating and amplifying rat bone marrow stromal cells in vitro, so as to lay a foundation for further study and utilization of the cells. 2. To study the feasibility and effectiveness of gene modification of bone marrow stromal cells by liposome transfection, and to study the application of bone marrow stromal cells in tissue engineering and gene therapy. [Methods Rat bone marrow stromal cells were isolated from lymphocytes and cultured with DMEM medium containing 20% fetal bovine serum for trypsin digestion and passage. Rat bone marrow stromal cells were amplified. The third passage BMSCs were used to transfect the green fluorescent protein gene into the bone marrow stromal cells by lipofectamine transfection, and then observed by fluorescence microscope. The number of cells expressing fluorescent protein was counted and the viability of transfected cells was detected by trypan blue exclusion test. [Results] 1 isolation and culture of rat BMSC Percoll lymphocyte isolate was used to centrifuge rat bone marrow with density gradient centrifugation. The cells were spherical mononuclear cells. The activity of Trypan blue stained cells was 99%. 24 hours after inoculation. Inverted microscope
【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2005
【分類(lèi)號(hào)】：R346

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本文編號(hào)：1438622