Simultaneous Detection of FLT3-ITD and NPM1Gene Mutations in Acute Myeloid Leukemia by Double PCR

本研究旨在建立一种同时筛查flT3-ITd突变和nPM1突变的检测方法。设计2对引物,分别扩增nPM1基因的外显子12和flT3基因的外显子14、内含子14、外显子15,以覆盖几乎所有已知突变位点。对双重PCr体系的反应程序和引物浓度比例进行优化,将双重PCr产物通过毛细管电泳分离,根据野生型产物和突变型产物的大小差异来判断突变存在与否并利用产物的峰面积对突变比例进行定量,并对突变阳性标本进行测序验证。结果表明,在93例标本中nPM1突变者17例(18.5%),flT3-ITd突变者15例(16.3%),nPM1突变和flT3-ITd突变双阳性6例。17例nPM1突变中7例M2、4例M4、5例M5、1例M6,其中10例男性、7例女性;有15例为A型,1例为b型,1例为nM型;有1例CMl急变为AMl的标本中带有nPM1基因A型突变。15例flT3-ITd阳性中1例M1、8例M2、2例M3、1例M4、3例M5,其中5例男性、10例女性。扩增产物测序结果进一步证明了该检测体系的准确性和可靠性。结论 :建立了同时筛查flT3-ITd突变和nPM1突变的检测体系,该体系以基因组dnA为模板,具有方便、快捷、准确、可定量的优点。Objective of this study was to establish a method for simultaneous detection of FLT3/ITD and NPM1 gene mutations in AML.A double PCR was firstly designed and optimized to amplify both exon 12 of NPM1 and exon 14-intron 14-exon 15 of FLT3,with the aim of detecting almost all reported mutations.After optimization,a touchdown PCR was chosen for the multiplex PCR procedure,with the primer concentrations of NPM1 and FLT3-ITD being 200 nmol/L and 152 nmol/L respectively.The PCR amplicons were separated by capillary electrophoresis and the presence of mutants was recognized by the size difference between the mutants and wild-type products.The areas of mutant peak and wild-type peak were used to calculate the mutant/wild-type ratio.All the positive mutated samples were confirmed by sequencing.The results showed that 17 patients with NPM1 mutation,15 patients with FLT3-ITD mutation,6 patients with both NPM1 and FLT3-ITD mutationts were found among 93 patents.7 patients with M2,4 patients with M4,5 patients with M5 and 1 patients with M6 were found out of 17 patients with NPM1 mutation,in which 10 patients were male and 7 patients were female,15 patients were with type A,1 patients was with type B and 1 patients was with type Nm,strinkingly 1 CML patient in blast crisis was found to carry a type A mutation.Among 15 patients with FLT3-ITD mutation 1 patient with M1,8 patients with M2,2 patients with M2,2 patients with M3,1 patient with M4,3 patients with M5 were found,in which 5 patients were male and 10 patients were lemale.Sequencing results further confirmed the accuracy and reliability of this method.It is concluded that a novel method with the ability to detect both FLT3-ITD and NPM1 mutations has been developed when genomic DNA was templated.This method is fast,easy,accurate and capable to calculate the mutant/wild-type ratio

海洋硫酸盐还原菌群处理烟气脱硫废水

针对高盐硫酸根废水的特性,从海底沉积物中富集得到1个硫酸盐还原菌菌群SRB-2,并研究了盐度、温度、pH值、碳源、硫酸根浓度和铁形态对其活性的影响.结果表明,SRB-2为嗜盐中温硫酸盐还原菌群,可以利用乙醇及乳酸为单一碳源,最佳生长温度为30～40℃,最佳生长pH值为7.4～8.3,SRB-2菌群能够在硫酸根浓度为5200mg/L或盐度为60g/L的条件下正常生长,还原铁粉对该菌群还原硫酸根的能力具有加强作用,而二价铁离子则抑制细菌活性.扫描电子显微镜及光学显微镜对菌体形态研究发现,在反应器填料表面,该菌群中有大量短竿状及螺旋状细菌黏附,而在底部,主要为表面覆盖大量黑色粘稠物质,由短竿状细菌组成,推测其可能是菌种SRB-2-64(GenBank序列号:EU167911)的堆积

海洋硫酸盐还原菌群处理烟气脱硫废水

针对高盐硫酸根废水的特性，从海底沉积物中富集得到1个硫酸盐还原菌菌群SRB．2，并研究了盐度、温度、pH值、碳源、硫酸根浓度和铁形态对其活性的影响．结果表明，SRB-2为嗜盐中温硫酸盐还原菌群，可以利用乙醇及乳酸为单一碳源，最佳生长温度为30～40℃，最佳生长pH值为7．4～8．3，SRB．2菌群能够在硫酸根浓度为5200mg／L或盐度为60g／L的条件下正常生长，还原铁粉对该菌群还原硫酸根的能力具有加强作用，而二价铁离子则抑制细菌活性．扫描电子显微镜及光学显微镜对菌体形态研究发现，在反应器填料表面，该菌群中有大量短竿状及螺旋状细菌黏附，而在底部，主要为表面覆盖大量黑色粘稠物质，由短竿状细菌组成，推测其可能是菌种SRB-2—64（GenBank序列号：EU167911）的堆积

Phenylhexyl isothiocyanate induces gene p16 demethylation by down-regulating DNA methyltransferases in U266 cells

目的研究异硫氰酸苯己酯(PHI)对骨髓瘤u266细胞株P16基因的去甲基化作用并探讨其作用机制。方法应用半定量逆转录-聚合酶链反应检测u266细胞经过PHI处理后dnA甲基转移酶dnMT1、dnMT3A和dnMT3b基因的MrnA的表达变化,用甲基化特异性聚合酶链反应检测PHI作用前后u266细胞株P16基因甲基化状态的变化。结果使用不同浓度的PHI处理u266细胞72H后,u266细胞表达dnMT1、dnMT3A、dnMT3bMrnA的水平明显降低并呈浓度依赖性。以不同浓度的PHI处理u266细胞10d后,P16基因的甲基化状态被逐渐逆转。结论 PHI可能通过抑制dnA甲基转移酶的表达水平诱导P16基因产生去甲基化,使失活的抑癌基因重新激活,诱导瘤细胞凋亡。Objective To study the effect of demethylation of P16 and discuss the mechanisms by phenylhexyl isothiocyanate(PHI) on Myeloma U266 cells.Methods The mRNA expression of DNA methyltransferase 1,DNA methyltransferase 3a and DNA methyltransferase 3b were measured by RT-PCR,Modified methylation specific PCR(MSP) was used to screen p16-M and p15-U mRNA on U266 cells treated with PHI.Results After 72 hours treatment with different concentration PHI,the mRNA expression of DNMT1,DNMT3a and DNMT3b was decreased and related to PHI concentration,After 10 days PHI treatment,P16 gene hypermethylation was reversed at 10 μmol/L concentration.Conclusion By decreasing the level of DNA methyltransferase,PHI can reverse hypermethylation of gene P16 and re-actived silenced P16 gene and induce myeloma cells apoptosis in U266 cells.厦门市科技计划项目(3502Z20104034

海洋硫酸盐还原菌群处理烟气脱硫废水

针对高盐硫酸根废水的特性，从海底沉积物中富集得到1个硫酸盐还原菌菌群SRB．2，并研究了盐度、温度、pH值、碳源、硫酸根浓度和铁形态对其活性的影响．结果表明，SRB-2为嗜盐中温硫酸盐还原菌群，可以利用乙醇及乳酸为单一碳源，最佳生长温度为30～40℃，最佳生长pH值为7．4～8．3，SRB．2菌群能够在硫酸根浓度为5200mg／L或盐度为60g／L的条件下正常生长，还原铁粉对该菌群还原硫酸根的能力具有加强作用，而二价铁离子则抑制细菌活性．扫描电子显微镜及光学显微镜对菌体形态研究发现，在反应器填料表面，该菌群中有大量短竿状及螺旋状细菌黏附，而在底部，主要为表面覆盖大量黑色粘稠物质，由短竿状细菌组成，推测其可能是菌种SRB-2—64（GenBank序列号：EU167911）的堆积

JUNO Sensitivity on Proton Decay p→νˉK+p\to \bar\nu K^+ Searches

The Jiangmen Underground Neutrino Observatory (JUNO) is a large liquid scintillator detector designed to explore many topics in fundamental physics. In this paper, the potential on searching for proton decay in $p\to \bar\nu K^+$ mode with JUNO is investigated.The kaon and its decay particles feature a clear three-fold coincidence signature that results in a high efficiency for identification. Moreover, the excellent energy resolution of JUNO permits to suppress the sizable background caused by other delayed signals. Based on these advantages, the detection efficiency for the proton decay via $p\to \bar\nu K^+$ is 36.9% with a background level of 0.2 events after 10 years of data taking. The estimated sensitivity based on 200 kton-years exposure is $9.6 \times 10^{33}$ years, competitive with the current best limits on the proton lifetime in this channel

JUNO sensitivity on proton decay p → ν K + searches*

The Jiangmen Underground Neutrino Observatory (JUNO) is a large liquid scintillator detector designed to explore many topics in fundamental physics. In this study, the potential of searching for proton decay in the $p\to \bar{\nu} K^+$ mode with JUNO is investigated. The kaon and its decay particles feature a clear three-fold coincidence signature that results in a high efficiency for identification. Moreover, the excellent energy resolution of JUNO permits suppression of the sizable background caused by other delayed signals. Based on these advantages, the detection efficiency for the proton decay via $p\to \bar{\nu} K^+$ is 36.9% ± 4.9% with a background level of $0.2\pm 0.05({\rm syst})\pm$$0.2({\rm stat})$ events after 10 years of data collection. The estimated sensitivity based on 200 kton-years of exposure is $9.6 \times 10^{33}$ years, which is competitive with the current best limits on the proton lifetime in this channel and complements the use of different detection technologies