【摘要】：人類(lèi)p53基因存在于17p13.1染色體上,編碼分子量為53kDa的核磷酸化蛋白,p53基因是迄今發(fā)現(xiàn)的與人類(lèi)腫瘤相關(guān)性最高的基因。P53基因突變及其表達(dá)產(chǎn)物在腫瘤發(fā)生時(shí)常伴隨發(fā)生,且p53表達(dá)者預(yù)后較差,提示其與腫瘤的發(fā)生密切相關(guān),檢測(cè)其基因突變株及表達(dá)產(chǎn)物有助于腫瘤的診斷。p53基因突變后導(dǎo)致蛋白的組化性質(zhì)發(fā)生改變,蛋白的穩(wěn)定性大大增加,可通過(guò)分子生物學(xué)方法、免疫組化法、血清學(xué)等多種方法分析。而免疫組化方法是目前臨床上對(duì)腫瘤,特別是疑難腫瘤分析檢測(cè)所采用的重要方法,可以利用它分析p53基因突變誘導(dǎo)表達(dá)的蛋白在腫瘤組織中的情況。 本實(shí)驗(yàn)利用本實(shí)驗(yàn)組構(gòu)建的PQE-p53表達(dá)質(zhì)粒,在大腸桿菌M15表達(dá)系統(tǒng)中進(jìn)行p53表達(dá)。建立p53蛋白在原核生物中的表達(dá)純化方法,制備純化的p53蛋白及相對(duì)抗體。目前雖然用于p53抗原檢測(cè)的單、多抗很多,但大多是進(jìn)口產(chǎn)品。所以基于以上情況我們嘗試制備出可用于檢測(cè)的單、多克隆抗體對(duì)腫瘤石蠟切片進(jìn)行免疫分析,與進(jìn)口產(chǎn)品其相比價(jià)格會(huì)大大降低,有助于腫瘤p53基因表達(dá)和突變的研究。
[Abstract]:Human p53 gene exists on the 17p13.1 chromosome, and the p53 gene encoding 53kDa is the most relevant gene. P53 gene mutation and its expression products often occur in tumorigenesis. The prognosis of the patients with p53 expression was poor, which suggested that p53 gene mutation and its expression products were closely related to the occurrence of tumor. The detection of p53 gene mutation would help to change the histochemical properties of the protein and increase the stability of the protein. It can be analyzed by molecular biology, immunohistochemistry, serology and so on. Immunohistochemical method is an important method for the analysis and detection of tumor, especially difficult tumor. It can be used to analyze the expression of p53 gene mutation induced protein in tumor tissue. In this experiment, p53 was expressed in E. coli M15 expression system by using the PQE-p53 expression plasmid constructed in our experimental group. The method of expression and purification of p53 protein in prokaryote was established to prepare purified p53 protein and relative antibody. Although currently used for the detection of p53 antigen, many polyclonal antibodies, but mostly imported products. Therefore, we try to prepare a single, polyclonal antibody that can be used for detection of tumor paraffin sections. Compared with the imported products, the price will be greatly reduced, which is helpful for the study of p53 gene expression and mutation.
【學(xué)位授予單位】：東北師范大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2006
【分類(lèi)號(hào)】：R392

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本文編號(hào)：2170959