A loop-mediated isothermal amplification （LAMP） assay was designed and evaluated for rapid de- tection of the toxic microalgae Alexandrium catenella and A. minutum, which can produce paralytic shellfish poisoning （...A loop-mediated isothermal amplification （LAMP） assay was designed and evaluated for rapid de- tection of the toxic microalgae Alexandrium catenella and A. minutum, which can produce paralytic shellfish poisoning （PSP）. Two sets of four specific primers targeting these two species were derived from the sequence of internal transcribed spacer （ITS） of ribosomal DNA. The method worked well in less than an hour under isothermal conditions of 65℃. LAMP specificity was validated in closely related algae as a comparison, suggesting the strict specificity of the LAMP primers. Two visual inspection approaches were feasible to interpret the positive or negative results. The detection lim- its of A. catenella and A. minutum samples using the LAMP assay were found to be 5.6 and 4.5 pg DNA, respectively. The sensitivity of this LAMP assay was 10 or 100-fold higher than Polymerase Chain Reaction （PCR） method in detecting the two microalgae. These characteristics of species specificity, sensitivity, and rapidity suggest that this method has the potentiality in the monitoring of red tide caused by A. catenella and A. minutum.展开更多
MicroRNAs(miRNAs) are an abundant class of conserved, non-coding small RNAs that play important role in gene regulation at post translational level. There have been no reports on the miRNAs of the Antarctic krill E up...MicroRNAs(miRNAs) are an abundant class of conserved, non-coding small RNAs that play important role in gene regulation at post translational level. There have been no reports on the miRNAs of the Antarctic krill E uphausia superba despite the species' crucial position in Antarctic food webs. Two small RNA libraries were constructed from eyestalk and muscle, subsequently, and deep sequencing analysis was performed to investigate and profile E. superba miRNAs. A total of 19 304 586 and 23 005 104 unique sequences were obtained from the eyestalk and muscle, respectively. After compared the small RNA sequences with the Rfam database, 12 342 039 and 7 907 477 reads in eyestalk and muscle were matched to the transcriptome sequence of E. superba. A total of 236 distinct miRNAs were identified after annotation to known animal miRNAs registered in miRBase 21. In both libraries, the most abundant known miRNA were miR-750 with 92 583 reads in muscle and miR-1304-3 p with 56 386 reads in eyestalk while the average value was less than 106, revealing a wide range of different expression levels. In addition, miR-277 a enriched in both libraries and may be involved in modulating Krebs cycle by targeting to Vimar. Differential expression analysis showed that 56 mature miRNAs were significantly up/down regulated according to expression fold change. It appeared that the expression of several abundant miRNAs maybe tissue-specific or tissue-bias. Notably, the expression pattern of miR-750 and miR-1 family, which was suggested as the crucial candidates, involved in muscle development. Taken together, this study provides the first miRNA profile of E. superba and some of these miRNAs were expected to play important role in immune response, reproduction, energy metabolism, and muscle development and so on. Thus, the results provides a reference for functional studies of miRNAs in E. superba.展开更多
基金The Science and Technology Commission of Shanghai Municipality under contract Nos 06235810108DZ1980802 and 10JC1418600a special research fund for the national non-profit institutes (East China Sea Fisheries Research Institute) under contract Nos 2007M22 and 2007Z01
文摘A loop-mediated isothermal amplification （LAMP） assay was designed and evaluated for rapid de- tection of the toxic microalgae Alexandrium catenella and A. minutum, which can produce paralytic shellfish poisoning （PSP）. Two sets of four specific primers targeting these two species were derived from the sequence of internal transcribed spacer （ITS） of ribosomal DNA. The method worked well in less than an hour under isothermal conditions of 65℃. LAMP specificity was validated in closely related algae as a comparison, suggesting the strict specificity of the LAMP primers. Two visual inspection approaches were feasible to interpret the positive or negative results. The detection lim- its of A. catenella and A. minutum samples using the LAMP assay were found to be 5.6 and 4.5 pg DNA, respectively. The sensitivity of this LAMP assay was 10 or 100-fold higher than Polymerase Chain Reaction （PCR） method in detecting the two microalgae. These characteristics of species specificity, sensitivity, and rapidity suggest that this method has the potentiality in the monitoring of red tide caused by A. catenella and A. minutum.
基金Supported by the Sub-project under National Science&Technology Support Plan(No.2013BAD13B03)the Special Fund for Agroscientific Research in the Public Interest(No.201203018)
文摘MicroRNAs(miRNAs) are an abundant class of conserved, non-coding small RNAs that play important role in gene regulation at post translational level. There have been no reports on the miRNAs of the Antarctic krill E uphausia superba despite the species' crucial position in Antarctic food webs. Two small RNA libraries were constructed from eyestalk and muscle, subsequently, and deep sequencing analysis was performed to investigate and profile E. superba miRNAs. A total of 19 304 586 and 23 005 104 unique sequences were obtained from the eyestalk and muscle, respectively. After compared the small RNA sequences with the Rfam database, 12 342 039 and 7 907 477 reads in eyestalk and muscle were matched to the transcriptome sequence of E. superba. A total of 236 distinct miRNAs were identified after annotation to known animal miRNAs registered in miRBase 21. In both libraries, the most abundant known miRNA were miR-750 with 92 583 reads in muscle and miR-1304-3 p with 56 386 reads in eyestalk while the average value was less than 106, revealing a wide range of different expression levels. In addition, miR-277 a enriched in both libraries and may be involved in modulating Krebs cycle by targeting to Vimar. Differential expression analysis showed that 56 mature miRNAs were significantly up/down regulated according to expression fold change. It appeared that the expression of several abundant miRNAs maybe tissue-specific or tissue-bias. Notably, the expression pattern of miR-750 and miR-1 family, which was suggested as the crucial candidates, involved in muscle development. Taken together, this study provides the first miRNA profile of E. superba and some of these miRNAs were expected to play important role in immune response, reproduction, energy metabolism, and muscle development and so on. Thus, the results provides a reference for functional studies of miRNAs in E. superba.
