熒光蛋白可以用于標(biāo)記蛋白，在特定激發(fā)光激發(fā)下會(huì)發(fā)出特定的熒光，廣泛應(yīng)用于活細(xì)胞實(shí)時(shí)觀察。熒光蛋白標(biāo)記結(jié)核分枝桿菌特定蛋白后，可以研究該蛋白的定位，測(cè)定結(jié)核分枝桿菌氧化還原水平，研究結(jié)核分枝桿菌在動(dòng)物細(xì)胞中的動(dòng)態(tài)變化等，有助于結(jié)核分枝桿菌免疫學(xué)領(lǐng)域的研究。

Figure 7. M.tb localizes to LC3-positive compartments. (A) GFPLC3expressing macrophages were infected with mCherry-M.tb (MOI 50

or 10, 6 h) and then fixed and observed by confocal microscopy. At high MOI there are GFP-LC3 rings surrounding (arrowhead) around mCherry-M.tb and co-localization indicated by yellow pixels (arrow) that are not seen in macrophages challenged at low MOI. (B) LAMP1 immunostaining of GFP-LC3 cell line macrophages infected with M.tb Edrman (MOI 50 or 10, 5 h) demonstrates co-localization of LC3 and lysosomal markers (arrowheads).

doi:10.1371/journal.pone.0027972.g007

Figure 2. Differences in expression of rv2390c9::GFP in Mtb present in vaccinated versus mock-treated mice. Erdman(rv2390c9::GFP, smyc9::mCherry) was inoculated into vaccinated or mock-treated C57BL/6J WT mice for up to 56 days. (A) shows 3D confocal images from a 14, 28, or 42 day infection. All bacteria are marked in red (smyc9::mCherry), reporter signal is shown in green (rv2390c9::GFP), nuclei are marked in grayscale (DAPI), and phalloidin staining of f-actin is shown in blue. Scale bar 10 mm. (B) shows quantification of the GFP/mm3 signal for each bacterium measured from multiple 3D confocal images, at the indicated time points. Each point on the graph represents a bacterium or a tightly clustered group of bacteria (mock-treated – filled symbols, vaccinated – open symbols). Horizontal lines mark the median value for each group. p-values were obtained with a Mann-Whitney statistical test.

Interactions between Na?ve and Infected Macrophages Reduce Mycobacterium tuberculosis Viability

應(yīng)用分枝桿菌過表達(dá)質(zhì)粒pMV261（游離型）、pMV361（整合型），構(gòu)建eGFP（綠色熒光）、mCherry（紅色熒光）熒光質(zhì)粒，使熒光蛋白在分枝桿菌內(nèi)持續(xù)表達(dá)，感染巨噬細(xì)胞或其他細(xì)胞，檢測(cè)感染效率、細(xì)胞自噬、細(xì)胞凋亡、結(jié)核分枝桿菌存活能力等作用；感染小鼠，檢測(cè)結(jié)核分枝桿菌定位；另外，熒光菌株還可用于肺外結(jié)核如腸結(jié)核、腦結(jié)核、骨結(jié)核等定位研究，助力于結(jié)核分枝桿菌致病機(jī)制的研究。

熒光菌株示例圖

注：圖A為恥垢分枝桿菌 eGFP、mCherry熒光菌株在藍(lán)光激發(fā)下發(fā)出綠色和紅色熒光；圖B為牛結(jié)核分枝桿菌BCG eGFP、mCherry熒光菌株在藍(lán)光激發(fā)下發(fā)出綠色和紅色熒光；圖C為結(jié)核分枝桿菌H37Ra eGFP、mCherry熒光菌株在藍(lán)光激發(fā)下發(fā)出綠色和紅色熒光

晶諾生物已有恥垢分枝桿菌、牛結(jié)核分枝桿菌BCG、結(jié)核分枝桿菌H37Rv、結(jié)核分枝桿菌H37Ra的紅色熒光（mCherry）、綠色熒光（eGFP）菌株，歡迎各位老師訂購(gòu)。正在研發(fā)藍(lán)色熒光（BFP）、黃色熒光（YFP）、近紅外（iRFP720）、熒光素酶等標(biāo)記的分枝桿菌菌株，請(qǐng)各位老師持續(xù)關(guān)注！

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