An assessment with assistance of DNA barcoding was conducted on green macroalgae in coastal zone around Qingdao, China, during the period of April- December, 2011. Three markers were applied in molecular discriminatio...An assessment with assistance of DNA barcoding was conducted on green macroalgae in coastal zone around Qingdao, China, during the period of April- December, 2011. Three markers were applied in molecular discrimination, including the plastid elongation factor tufA gene, the internal transcribed spacer(ITS) region of the ribosomal cistron and rubisco large subunit gene 3' regions(rbcL-3P). DNA barcoding discriminated 8 species, excluding species of genus Cladophora and Bryopsis due to failures in amplification. We ascertained and corrected 4 species identified by morphological methods for effectively assisting the classification. The gene tufA presented more advantages as an appropriate DNA marker with the strongest amplification success rate and species discrimination power than the other two genes. The poorest sequencing success largely handicapped the application of ITS. Samples identified by tufA and rbcL as Ulva flexuosa were clustered into the clade of U. prolifera by ITS in the neighbor-joining tree. Confusion with discrimination of the complex of U. linza, U. procera and U. prolifera(as the LPP complex) still existed for the three DNA markers. Based on our results, rbcL is recommended as a preferred marker for assisting tufA to discriminate green macroalgae. In distinguishing green-tide-forming Ulva species, the free-floating sample collected from the green tide in 2011 was proved to be identical with U. prolifera in Yellow Sea for ITS and rbcL genes. This study presents a preliminary survey of green macroalgae distributed in the coastal area around Qingdao, and proves that DNA barcoding is a powerful tool for taxonomy of green macroalgae.展开更多
目的探讨非小细胞肺癌(NSCLC)侵袭性边缘(IM)中CD8^(+)组织驻留记忆T细胞(CD8^(+)Trm)的表达和二维空间分布及其对复发的影响。方法回顾性收集2014-01-01-2018-12-31在山东省肿瘤医院行根治性手术的104例局部晚期NSCLC患者资料,并随访...目的探讨非小细胞肺癌(NSCLC)侵袭性边缘(IM)中CD8^(+)组织驻留记忆T细胞(CD8^(+)Trm)的表达和二维空间分布及其对复发的影响。方法回顾性收集2014-01-01-2018-12-31在山东省肿瘤医院行根治性手术的104例局部晚期NSCLC患者资料,并随访无复发生存期(RFS)。使用全自动组织微阵列制作仪(TMA Grand Master,3DHISTECH Ltd.)构建组织芯片,并利用多色免疫荧光技术精准量化CD8、CD8^(+)Trm的密度及接近度(20/30/40μm)。使用X-Tile确定截断值,将患者分为高/低密度/接近度组,Kaplan-Meier法及log-rank检验评估RFS差异。Cox比例风险回归模型确定关键复发因子。结果在局部晚期NSCLC患者的IM区域中,CD8^(+)Trm约占CD8的29.0%,CD8与CD8^(+)Trm高浸润患者分别为17(16.4%)和60例(57.7%),CD8及CD8^(+)Trm高接近度(20μm)患者分别为76(73.1%)和43例(41.4%)。多因素Cox分析显示,CD8及CD8^(+)Trm细胞的密度均非局部晚期NSCLC患者预后评估的独立风险因素,χ^(2)=1.578,P=0.209;χ^(2)=0.483,P=0.487。整体CD8的接近度亦与复发无关,但CD8^(+)Trm高接近度组(30μm)患者RFS高于低接近度组患者,P=0.008,HR=0.41,95%CI:0.22~0.79。结论IM区域浸润的CD8^(+)Trm与肿瘤细胞的频繁相互作用可能促进肿瘤免疫,降低局部晚期NSCLC复发风险。展开更多
基金supported by the Public Science and Technology Research Funds Projects of Ocean(201105021 and 201305030)National Natural Science Foundation China(41276137)+1 种基金Shandong Provincial Natural Science Foundation,China(ZR2011CM 018)Qingdao Municipal Science and Technology Program,China(09-2-5-3-hy)
文摘An assessment with assistance of DNA barcoding was conducted on green macroalgae in coastal zone around Qingdao, China, during the period of April- December, 2011. Three markers were applied in molecular discrimination, including the plastid elongation factor tufA gene, the internal transcribed spacer(ITS) region of the ribosomal cistron and rubisco large subunit gene 3' regions(rbcL-3P). DNA barcoding discriminated 8 species, excluding species of genus Cladophora and Bryopsis due to failures in amplification. We ascertained and corrected 4 species identified by morphological methods for effectively assisting the classification. The gene tufA presented more advantages as an appropriate DNA marker with the strongest amplification success rate and species discrimination power than the other two genes. The poorest sequencing success largely handicapped the application of ITS. Samples identified by tufA and rbcL as Ulva flexuosa were clustered into the clade of U. prolifera by ITS in the neighbor-joining tree. Confusion with discrimination of the complex of U. linza, U. procera and U. prolifera(as the LPP complex) still existed for the three DNA markers. Based on our results, rbcL is recommended as a preferred marker for assisting tufA to discriminate green macroalgae. In distinguishing green-tide-forming Ulva species, the free-floating sample collected from the green tide in 2011 was proved to be identical with U. prolifera in Yellow Sea for ITS and rbcL genes. This study presents a preliminary survey of green macroalgae distributed in the coastal area around Qingdao, and proves that DNA barcoding is a powerful tool for taxonomy of green macroalgae.
文摘目的探讨非小细胞肺癌(NSCLC)侵袭性边缘(IM)中CD8^(+)组织驻留记忆T细胞(CD8^(+)Trm)的表达和二维空间分布及其对复发的影响。方法回顾性收集2014-01-01-2018-12-31在山东省肿瘤医院行根治性手术的104例局部晚期NSCLC患者资料,并随访无复发生存期(RFS)。使用全自动组织微阵列制作仪(TMA Grand Master,3DHISTECH Ltd.)构建组织芯片,并利用多色免疫荧光技术精准量化CD8、CD8^(+)Trm的密度及接近度(20/30/40μm)。使用X-Tile确定截断值,将患者分为高/低密度/接近度组,Kaplan-Meier法及log-rank检验评估RFS差异。Cox比例风险回归模型确定关键复发因子。结果在局部晚期NSCLC患者的IM区域中,CD8^(+)Trm约占CD8的29.0%,CD8与CD8^(+)Trm高浸润患者分别为17(16.4%)和60例(57.7%),CD8及CD8^(+)Trm高接近度(20μm)患者分别为76(73.1%)和43例(41.4%)。多因素Cox分析显示,CD8及CD8^(+)Trm细胞的密度均非局部晚期NSCLC患者预后评估的独立风险因素,χ^(2)=1.578,P=0.209;χ^(2)=0.483,P=0.487。整体CD8的接近度亦与复发无关,但CD8^(+)Trm高接近度组(30μm)患者RFS高于低接近度组患者,P=0.008,HR=0.41,95%CI:0.22~0.79。结论IM区域浸润的CD8^(+)Trm与肿瘤细胞的频繁相互作用可能促进肿瘤免疫,降低局部晚期NSCLC复发风险。
