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  • orfs  (ATCC)
    85
    ATCC orfs
    Representative examples of <t>mRNA</t> decay patterns for selected operons from B. cereus ATCC 10897 . For each operon, the upper graph displays the coverage at t0 plotted from the start to the end of the operon, while the lower graph shows, for each single nucleotide position, the log2 ratio of the sequence coverage at different time-points following rifampicin addition (red, t2.5, 2.5 minutes; green, t5, 5 minutes; blue, t10, 10 minutes), relative to the sequence coverage at t0. (a) Operons demonstrating examples of 5' to 3' direction of mRNA decay, that is, the sequence coverage for bases near the 5' end of each ORF in the mRNA drops faster than for bases at the 3' end of <t>ORFs.</t> (b) Operons demonstrating examples of 3' to 5' direction of mRNA decay. The seemingly anomalous pattern for the 2.5 minute or 5 minute time-points in the 5' end of the TU_4695703-4696150_R transcript (green graph, leftmost panel) may be due to technical noise. (c) Operons showing various complex types of decay patterns, where specific blocks of the polycistronic mRNA are degraded faster, or show a pattern of faster decay at both 5' and 3' ends relative to other parts of the mRNA.
    Orfs, supplied by ATCC, used in various techniques. Bioz Stars score: 85/100, based on 501 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Biotechnology Information orf finder
    Schematic diagram for the evaluation of promoter activity. ( a ) Outline of the construction of transformation vectors and transformations. After predicting putative <t>ORF</t> positions 32 , upstream regions of the <t>ORFs</t> were determined as potential promoter regions. Potential promoter regions amplified by PCR were used to construct the transformation vectors. The double-cassette vector containing the reporter gene egfp driven by each tested promoter and the antibiotic-resistant gene Sh ble driven by the promoter region of the fucoxanthin chlorophyll a / c -binding protein (FCP) A-1A gene derived from Cyl. fusiformis (termed CffcpA pro.) were constructed. ( b ) Assessment of promoter activity. Promoter activity was determined by averaging the ratios of egfp mRNA transcript levels to those of Sh ble mRNA transcripts in ten transformants to minimize the effects of copy numbers on the expression of transgenes. These transformants were also used to investigate eGFP protein expression patterns. CffcpA ter.: terminator region of the FCP A-1A gene derived from Cyl. fusiformis . The structure of the ClorDNA genome was modified from Tomaru et al. 32 . *For the transformation vector of the nitrate reductase gene promoter, we used pNICgfp 18 ( Supplementary Fig. 5a ).
    Orf Finder, supplied by Biotechnology Information, used in various techniques. Bioz Stars score: 91/100, based on 316 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Thermo Fisher orfs
    Creating and screening large CASTLING libraries. ( a ) Three libraries with different numbers of collected clones were generated from SIC pools combining either 2 or 30 recombineering reactions to investigate the minimum effort for a proteome-wide (design: 5,940 <t>ORFs,</t> oligonucleotide pool D, Supplementary Table 2 ) CASTLING library (details in Methods). ( b ) Venn diagram of genotypes recovered in each of the three libraries; all libraries combined tagged 4,516 different ORFs. ( c ) Genotype diversity in each of the three libraries, shared between them, or after their combination. ( d ) Proteome profiling by fluorescence intensity of a non-exhaustive mNeonGreen tag library (library #1.1, Figure 4c , Supplementary Table 2 ) using FACS. After enriching the fluorescent sub-population of the library and determining the fold-enrichment of each genotype by NGS, this sub-population was sorted into eight bins according to fluorescent intensity. Analysis of each bin by Anchor-Seq and on-site nanopore sequencing allowed the assignment of an expected protein abundance for each genotype. ( e ) Pairwise comparisons between fluorescence intensity estimates calculated from genotype distribution across all bins (Methods, Equation 2 ; this study denoted as BUC) and protein abundances reported by selected genome-scale experiments 4 , 39 , 40 normalized to molecules per cell 38 . Outliers (orange) were determined based on the comparison to a <t>GFP</t> tag flow cytometry study 39 . Spearman correlation coefficients (ρ) are given. Marginal lines indicate abundance estimates only present in the respective study but missing in the other. ( f ) Comparison of Spearman correlation coefficients between studies either considering their overlap in detected ORFs or only the overlap with the 435 ORFs we could detect in this experiment. A Pearson correlation coefficient (r) is given. ( g ) Eight genes that had not been characterized in other genome-scale experiments 38 were tagged individually to verify whether fluorescence intensity corresponded with their predicted characterization by FACS. Same exposure time for all fluorescent microscopy images except for Ybr196c-a which was imaged at 10% excitation; scale bar 10 µm. Panels b, c and e: Source data are provided as a Source Data file.
    Orfs, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 94/100, based on 3568 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    OriGene gfp tagged orf
    Rsu-1 levels in HCC. A) Graphical representation of the number of HCC cases positive, negative or moderately positive for Rsu-1 in HCC tissue array (24 cases/48 cores). B) Protein levels of Rsu-1 in HCC cell lines compared to human hepatocytes (HH). Most of the HCC cell lines show decrease in Rsu-1 protein. C) Successful overexpression of <t>Rsu-GFP</t> (fusion protein) in Hep3B cell line. GFP tagged <t>ORF</t> clone of Homo sapiens Rsu-1 (#RG203334, Origene) was transfected into Hep3B cell line and analyzed for Rsu-1 48 h after transfection. Since it is a GFP fused protein, the MW of Rsu-1 is ~fifty-five kd instead of twenty-nine kd (MW of GFP is ~twenty-six kd). D) GFP-Rsu-1 fusion protein associates with PINCH inside the cell. Overexpression of GFP-Rsu-1 in Hep3B cell line leads to association of GFP-Rsu-1 with PINCH. GFP was immunoprecipitated 48 h after transfection. GFP precipitates were probed with either GFP or PINCH. Presence of PINCH in GFP precipitates shows association of GFP-Rsu-1with PINCH.
    Gfp Tagged Orf, supplied by OriGene, used in various techniques. Bioz Stars score: 88/100, based on 216 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    RayBiotech vascular endothelial growth factor vegf
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
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    88
    Biotechnology Information orf finder tool
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Orf Finder Tool, supplied by Biotechnology Information, used in various techniques. Bioz Stars score: 88/100, based on 69 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    OriGene myc ddk tagged orf
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Myc Ddk Tagged Orf, supplied by OriGene, used in various techniques. Bioz Stars score: 88/100, based on 97 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Biotechnology Information open reading frames orfs
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Open Reading Frames Orfs, supplied by Biotechnology Information, used in various techniques. Bioz Stars score: 91/100, based on 117 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    OriGene human cdna orf
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Human Cdna Orf, supplied by OriGene, used in various techniques. Bioz Stars score: 88/100, based on 101 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Biotechnology Information orf finder software
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Orf Finder Software, supplied by Biotechnology Information, used in various techniques. Bioz Stars score: 88/100, based on 29 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    4Gene varicella zoster virus open reading frame 4 gene
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Varicella Zoster Virus Open Reading Frame 4 Gene, supplied by 4Gene, used in various techniques. Bioz Stars score: 88/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Thermo Fisher yeast orf collection
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Yeast Orf Collection, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 110 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Illumina Inc arg like orfs
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Arg Like Orfs, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 93/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Thermo Fisher orf rnai library
    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, <t>HUVEC</t> were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and <t>VEGF</t> dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).
    Orf Rnai Library, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Promega firefly luciferase orf
    Experimental validation of target gene predictions. Validation of putative cell cycle relevant target genes for microRNAs miR-17 ( A ), miR-20a ( B ), and miR-106b ( C ) in HEK293T-cells. 3′-UTR fragments from putative targets CCND1, CCND2, E2F1, E2F3, CDKN1A, MAPK9, PTEN, RB1, RBL1, RBL2, and WEE1 were cloned at the 3′-end of the Firefly <t>ORF</t> in <t>Firefly/Renilla</t> dual reporter vector pmirGLO. To test the influence of endogenous microRNAs, pmirGLO and pmirGLO/3′-UTR were each transfected into HEK293T-cells. Normalized Firefly -activities were compared to those of pairwise co-transfections of these vectors with the microRNA mimic of interest (miR-17, miR-20a, miR-106b, also including an unspecific mimic negative control) to test for (i) unspecific effects of the given microRNA-mimic on Firefly/Renilla per se , (ii) effects of endogenous HEK293T microRNAs (iii) for validation of the particular target prediction. Dark grey columns show normalized Firefly activities from pmirGLO/3′-UTR transfections and pmirGLO + mimics co-transfections, light grey columns those from pmirGLO/3′-UTR + mimics co-transfections. Percent reductions of Firefly activities from co-transfections of pmirGLO/3′-UTR + mimic compared to pmirGLO + mimic are given as mean values from 2 biological experiments, each consisting of 4 technical replicates. Error bars represent standard deviations and statistical significancies (Student's t -test, unpaired, ***: p≤0.001, **: p≤0.01) are given. Effects of the unspecific negative control on pmirGLO and pmirGLO/3′-UTR vectors are shown separately in Fig.S2 . Except for E2F3 and MAPK9, all predicted proteins could be validated as statistically significant targets for all three microRNAs tested, although microRNA influence varied from strong (i.e. RBL2) to moderate (i.e. WEE1).
    Firefly Luciferase Orf, supplied by Promega, used in various techniques. Bioz Stars score: 89/100, based on 162 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    ASKA Pharmaceutical e coli k 12 orf archive
    Schematic overview of the high-throughput screening (HTS) for the determination of the bacterial interactive metabolites using the  E. coli  K-12 Keio collection.
    E Coli K 12 Orf Archive, supplied by ASKA Pharmaceutical, used in various techniques. Bioz Stars score: 88/100, based on 42 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Unigene orfs
    Flow chart of <t>ORF</t> prediction process whereby the translated Texas synganglion transcriptome <t>ORFs</t> are routed through TMHMM and GPCRPred to identify candidate GPCR-encoding sequences. The transcriptome sequences were translated into all 6 possible ORFs
    Orfs, supplied by Unigene, used in various techniques. Bioz Stars score: 92/100, based on 39 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    Promega rluc orf
    Flow chart of <t>ORF</t> prediction process whereby the translated Texas synganglion transcriptome <t>ORFs</t> are routed through TMHMM and GPCRPred to identify candidate GPCR-encoding sequences. The transcriptome sequences were translated into all 6 possible ORFs
    Rluc Orf, supplied by Promega, used in various techniques. Bioz Stars score: 85/100, based on 49 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Promega firefly luciferase open reading frame
    Flow chart of <t>ORF</t> prediction process whereby the translated Texas synganglion transcriptome <t>ORFs</t> are routed through TMHMM and GPCRPred to identify candidate GPCR-encoding sequences. The transcriptome sequences were translated into all 6 possible ORFs
    Firefly Luciferase Open Reading Frame, supplied by Promega, used in various techniques. Bioz Stars score: 88/100, based on 138 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    89
    Promega flexi orf
    Flow chart of <t>ORF</t> prediction process whereby the translated Texas synganglion transcriptome <t>ORFs</t> are routed through TMHMM and GPCRPred to identify candidate GPCR-encoding sequences. The transcriptome sequences were translated into all 6 possible ORFs
    Flexi Orf, supplied by Promega, used in various techniques. Bioz Stars score: 89/100, based on 134 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Genecopoeia orf cdna lentiviral particles
    Flow chart of <t>ORF</t> prediction process whereby the translated Texas synganglion transcriptome <t>ORFs</t> are routed through TMHMM and GPCRPred to identify candidate GPCR-encoding sequences. The transcriptome sequences were translated into all 6 possible ORFs
    Orf Cdna Lentiviral Particles, supplied by Genecopoeia, used in various techniques. Bioz Stars score: 88/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Representative examples of mRNA decay patterns for selected operons from B. cereus ATCC 10897 . For each operon, the upper graph displays the coverage at t0 plotted from the start to the end of the operon, while the lower graph shows, for each single nucleotide position, the log2 ratio of the sequence coverage at different time-points following rifampicin addition (red, t2.5, 2.5 minutes; green, t5, 5 minutes; blue, t10, 10 minutes), relative to the sequence coverage at t0. (a) Operons demonstrating examples of 5' to 3' direction of mRNA decay, that is, the sequence coverage for bases near the 5' end of each ORF in the mRNA drops faster than for bases at the 3' end of ORFs. (b) Operons demonstrating examples of 3' to 5' direction of mRNA decay. The seemingly anomalous pattern for the 2.5 minute or 5 minute time-points in the 5' end of the TU_4695703-4696150_R transcript (green graph, leftmost panel) may be due to technical noise. (c) Operons showing various complex types of decay patterns, where specific blocks of the polycistronic mRNA are degraded faster, or show a pattern of faster decay at both 5' and 3' ends relative to other parts of the mRNA.

    Journal: Genome Biology

    Article Title: Global mRNA decay analysis at single nucleotide resolution reveals segmental and positional degradation patterns in a Gram-positive bacterium

    doi: 10.1186/gb-2012-13-4-r30

    Figure Lengend Snippet: Representative examples of mRNA decay patterns for selected operons from B. cereus ATCC 10897 . For each operon, the upper graph displays the coverage at t0 plotted from the start to the end of the operon, while the lower graph shows, for each single nucleotide position, the log2 ratio of the sequence coverage at different time-points following rifampicin addition (red, t2.5, 2.5 minutes; green, t5, 5 minutes; blue, t10, 10 minutes), relative to the sequence coverage at t0. (a) Operons demonstrating examples of 5' to 3' direction of mRNA decay, that is, the sequence coverage for bases near the 5' end of each ORF in the mRNA drops faster than for bases at the 3' end of ORFs. (b) Operons demonstrating examples of 3' to 5' direction of mRNA decay. The seemingly anomalous pattern for the 2.5 minute or 5 minute time-points in the 5' end of the TU_4695703-4696150_R transcript (green graph, leftmost panel) may be due to technical noise. (c) Operons showing various complex types of decay patterns, where specific blocks of the polycistronic mRNA are degraded faster, or show a pattern of faster decay at both 5' and 3' ends relative to other parts of the mRNA.

    Article Snippet: Employing B. cereus as a model, we provide genome-wide operon structure predictions for two B. cereus strains, mRNA half-lives for more than 2,700 ORFs, and mRNA degradation patterns at single nucleotide resolution for more than 500 operons in B. cereus ATCC 10987, a sequenced (closed) model strain that maps to a B. cereus group phylogenetic cluster that also encompasses B. anthracis .

    Techniques: Sequencing

    Schematic diagram for the evaluation of promoter activity. ( a ) Outline of the construction of transformation vectors and transformations. After predicting putative ORF positions 32 , upstream regions of the ORFs were determined as potential promoter regions. Potential promoter regions amplified by PCR were used to construct the transformation vectors. The double-cassette vector containing the reporter gene egfp driven by each tested promoter and the antibiotic-resistant gene Sh ble driven by the promoter region of the fucoxanthin chlorophyll a / c -binding protein (FCP) A-1A gene derived from Cyl. fusiformis (termed CffcpA pro.) were constructed. ( b ) Assessment of promoter activity. Promoter activity was determined by averaging the ratios of egfp mRNA transcript levels to those of Sh ble mRNA transcripts in ten transformants to minimize the effects of copy numbers on the expression of transgenes. These transformants were also used to investigate eGFP protein expression patterns. CffcpA ter.: terminator region of the FCP A-1A gene derived from Cyl. fusiformis . The structure of the ClorDNA genome was modified from Tomaru et al. 32 . *For the transformation vector of the nitrate reductase gene promoter, we used pNICgfp 18 ( Supplementary Fig. 5a ).

    Journal: Scientific Reports

    Article Title: Characterization of marine diatom-infecting virus promoters in the model diatom Phaeodactylum tricornutum

    doi: 10.1038/srep18708

    Figure Lengend Snippet: Schematic diagram for the evaluation of promoter activity. ( a ) Outline of the construction of transformation vectors and transformations. After predicting putative ORF positions 32 , upstream regions of the ORFs were determined as potential promoter regions. Potential promoter regions amplified by PCR were used to construct the transformation vectors. The double-cassette vector containing the reporter gene egfp driven by each tested promoter and the antibiotic-resistant gene Sh ble driven by the promoter region of the fucoxanthin chlorophyll a / c -binding protein (FCP) A-1A gene derived from Cyl. fusiformis (termed CffcpA pro.) were constructed. ( b ) Assessment of promoter activity. Promoter activity was determined by averaging the ratios of egfp mRNA transcript levels to those of Sh ble mRNA transcripts in ten transformants to minimize the effects of copy numbers on the expression of transgenes. These transformants were also used to investigate eGFP protein expression patterns. CffcpA ter.: terminator region of the FCP A-1A gene derived from Cyl. fusiformis . The structure of the ClorDNA genome was modified from Tomaru et al. 32 . *For the transformation vector of the nitrate reductase gene promoter, we used pNICgfp 18 ( Supplementary Fig. 5a ).

    Article Snippet: The ORFs of the two DIVs were identified using an ORF finder (National Center for Biotechnology Information, http://www.ncbi.nlm.nih.gov/gorf/ ).

    Techniques: Activity Assay, Transformation Assay, Amplification, Polymerase Chain Reaction, Construct, Plasmid Preparation, Binding Assay, Derivative Assay, Expressing, Modification

    ( A ) Schematic diagram of the genome organization of Botrytis cinerea hypovirus 1 (BcHV1) and Botrytis cinerea fusarivirus 1 (BcFV1). The coding strand of BcHV1 is 10,214 nt long and contains a large ORF encoding a polyprotein of 2964 aa, possessing conserved domains: Prot, papain-like protease; UGT, UDP glucose/sterol glucosyltransferase; RdRp, RNA-dependent RNA polymerase; Hel, viral helicase superfamily. The coding strand of BcFV1 is 8411 nt in length and contains two major ORFs, ORF1 and ORF2, encoding two polypeptides of 1644 aa and 715aa, respectively. ORF1-encoded polypeptide possesses two conserved domains, RdRp and Hel. ( B ) Northern blotting detection of BcHV1 and BcFV1 dsRNAs extracted from the mycelium of B. cinerea strain HBtom-372. Marker, λ-Hind III digest DNA marker.

    Journal: Viruses

    Article Title: Two Novel Hypovirulence-Associated Mycoviruses in the Phytopathogenic Fungus Botrytis cinerea: Molecular Characterization and Suppression of Infection Cushion Formation

    doi: 10.3390/v10050254

    Figure Lengend Snippet: ( A ) Schematic diagram of the genome organization of Botrytis cinerea hypovirus 1 (BcHV1) and Botrytis cinerea fusarivirus 1 (BcFV1). The coding strand of BcHV1 is 10,214 nt long and contains a large ORF encoding a polyprotein of 2964 aa, possessing conserved domains: Prot, papain-like protease; UGT, UDP glucose/sterol glucosyltransferase; RdRp, RNA-dependent RNA polymerase; Hel, viral helicase superfamily. The coding strand of BcFV1 is 8411 nt in length and contains two major ORFs, ORF1 and ORF2, encoding two polypeptides of 1644 aa and 715aa, respectively. ORF1-encoded polypeptide possesses two conserved domains, RdRp and Hel. ( B ) Northern blotting detection of BcHV1 and BcFV1 dsRNAs extracted from the mycelium of B. cinerea strain HBtom-372. Marker, λ-Hind III digest DNA marker.

    Article Snippet: Nucleotide Sequences and Amino Acid Residues Sequences Analysis ORFs in the full-length cDNA sequences of the dsRNAs in strain HBtom-372 of B. cinerea were deduced using the ORF Finder program in the website of the National Center for Biotechnology Information (NCBI, http://www.ncbi.nlm.nih.gov/gorf/gorf.html ).

    Techniques: Northern Blot, Marker

    Differential transcription of the genes for PTS Enzyme II and ABC transporters following growth of S. mutans UA159 in biofilm. Microarray results are presented as vertical bars. The numerical ORF designation (as presented in NCBI genome database) for

    Journal: Molecular oral microbiology

    Article Title: A Novel PTS of Streptococcus mutans is Responsible for Transport of Carbohydrates with ?-1,3 linkage

    doi: 10.1111/omi.12009

    Figure Lengend Snippet: Differential transcription of the genes for PTS Enzyme II and ABC transporters following growth of S. mutans UA159 in biofilm. Microarray results are presented as vertical bars. The numerical ORF designation (as presented in NCBI genome database) for

    Article Snippet: The OBio locus was analyzed using two ORF-calling programs, Glimmer ( , ) originally used for annotation of the S. mutans UA159 genome database and the National Center for Biotechnology Information ORF finder ( ).

    Techniques: Microarray

    Creating and screening large CASTLING libraries. ( a ) Three libraries with different numbers of collected clones were generated from SIC pools combining either 2 or 30 recombineering reactions to investigate the minimum effort for a proteome-wide (design: 5,940 ORFs, oligonucleotide pool D, Supplementary Table 2 ) CASTLING library (details in Methods). ( b ) Venn diagram of genotypes recovered in each of the three libraries; all libraries combined tagged 4,516 different ORFs. ( c ) Genotype diversity in each of the three libraries, shared between them, or after their combination. ( d ) Proteome profiling by fluorescence intensity of a non-exhaustive mNeonGreen tag library (library #1.1, Figure 4c , Supplementary Table 2 ) using FACS. After enriching the fluorescent sub-population of the library and determining the fold-enrichment of each genotype by NGS, this sub-population was sorted into eight bins according to fluorescent intensity. Analysis of each bin by Anchor-Seq and on-site nanopore sequencing allowed the assignment of an expected protein abundance for each genotype. ( e ) Pairwise comparisons between fluorescence intensity estimates calculated from genotype distribution across all bins (Methods, Equation 2 ; this study denoted as BUC) and protein abundances reported by selected genome-scale experiments 4 , 39 , 40 normalized to molecules per cell 38 . Outliers (orange) were determined based on the comparison to a GFP tag flow cytometry study 39 . Spearman correlation coefficients (ρ) are given. Marginal lines indicate abundance estimates only present in the respective study but missing in the other. ( f ) Comparison of Spearman correlation coefficients between studies either considering their overlap in detected ORFs or only the overlap with the 435 ORFs we could detect in this experiment. A Pearson correlation coefficient (r) is given. ( g ) Eight genes that had not been characterized in other genome-scale experiments 38 were tagged individually to verify whether fluorescence intensity corresponded with their predicted characterization by FACS. Same exposure time for all fluorescent microscopy images except for Ybr196c-a which was imaged at 10% excitation; scale bar 10 µm. Panels b, c and e: Source data are provided as a Source Data file.

    Journal: bioRxiv

    Article Title: Pooled clone collections by multiplexed CRISPR-Cas12a-assisted gene tagging in yeast

    doi: 10.1101/476804

    Figure Lengend Snippet: Creating and screening large CASTLING libraries. ( a ) Three libraries with different numbers of collected clones were generated from SIC pools combining either 2 or 30 recombineering reactions to investigate the minimum effort for a proteome-wide (design: 5,940 ORFs, oligonucleotide pool D, Supplementary Table 2 ) CASTLING library (details in Methods). ( b ) Venn diagram of genotypes recovered in each of the three libraries; all libraries combined tagged 4,516 different ORFs. ( c ) Genotype diversity in each of the three libraries, shared between them, or after their combination. ( d ) Proteome profiling by fluorescence intensity of a non-exhaustive mNeonGreen tag library (library #1.1, Figure 4c , Supplementary Table 2 ) using FACS. After enriching the fluorescent sub-population of the library and determining the fold-enrichment of each genotype by NGS, this sub-population was sorted into eight bins according to fluorescent intensity. Analysis of each bin by Anchor-Seq and on-site nanopore sequencing allowed the assignment of an expected protein abundance for each genotype. ( e ) Pairwise comparisons between fluorescence intensity estimates calculated from genotype distribution across all bins (Methods, Equation 2 ; this study denoted as BUC) and protein abundances reported by selected genome-scale experiments 4 , 39 , 40 normalized to molecules per cell 38 . Outliers (orange) were determined based on the comparison to a GFP tag flow cytometry study 39 . Spearman correlation coefficients (ρ) are given. Marginal lines indicate abundance estimates only present in the respective study but missing in the other. ( f ) Comparison of Spearman correlation coefficients between studies either considering their overlap in detected ORFs or only the overlap with the 435 ORFs we could detect in this experiment. A Pearson correlation coefficient (r) is given. ( g ) Eight genes that had not been characterized in other genome-scale experiments 38 were tagged individually to verify whether fluorescence intensity corresponded with their predicted characterization by FACS. Same exposure time for all fluorescent microscopy images except for Ybr196c-a which was imaged at 10% excitation; scale bar 10 µm. Panels b, c and e: Source data are provided as a Source Data file.

    Article Snippet: This would exceed available genome-wide tagging collection, e.g. the C-GFP collection with 4,159 ORFs (Thermo Fisher), the TAP-tag collection with 4,247 ORFs (Dharmacon) or our tandem fluorescent timer collection with 4,081 ORFs .

    Techniques: Clone Assay, Generated, Fluorescence, FACS, Next-Generation Sequencing, Nanopore Sequencing, Flow Cytometry, Microscopy

    CASTLING for tagging 215 nuclear proteins with a green fluorescent protein. ( a ) Three oligonucleotide pools of the same design (1,577 sequences, Supplementary Table 1 ) were used to create four tag libraries by CASTLING in duplicate sampling the indicated amount of starting material for PCR. ( b ) Detected oligonucleotide sequences of the design after PCR amplification (blue), self-integrating cassette (SIC) assembly (green) and in the final library (orange); oligonucleotides with copy number estimates (unique UMI counts) in the lowest quartile (lower 25%) are shown in light shade. ( c ) Same as (b), but evaluated in terms of open reading frames (ORFs) represented by the oligonucleotides or SICs. ( d ) Copy number of PCR amplicons recovered (red) or lost (blue) after recombineering; black horizontal lines indicate median UMI counts. ( e ) Pearson’s pairwise correlation of oligonucleotide or SIC copy number between replicates after PCR or rolling-circle amplification (RCA) respectively; n.s., not significant (p > 0.05). ( f ) Kernel density estimates of copy number in replicate 1a as normalized to the median copy number observed in the oligonucleotide pool (before recombineering) and after recombineering into the SIC pool (left panel); the distribution of fold-changes (right panel) highlights two frequency ranges: [0.1–0.9], i.e. 80% of SICs, and [0.25–0.75], i.e. 50% of SICs. ( g ) Representative fluorescence microscopy images of cells displaying nuclear, diffuse non-nuclear (asterisks), or no mNeonGreen fluorescence (arrows); scale bar 5 µm. ( h ) Quantification of fluorescence localization in > 1,000 cells in each replicate. ( i ) Recurrence of off-target events as revealed by Anchor-Seq across all library replicates and all genomic loci (left panel); the fraction of cells with SICs integrated at off-target sites (blue) within each clone population (red) is shown (right panel, axis trimmed). Panels b–i: Source data are provided as a Source Data file.

    Journal: bioRxiv

    Article Title: Pooled clone collections by multiplexed CRISPR-Cas12a-assisted gene tagging in yeast

    doi: 10.1101/476804

    Figure Lengend Snippet: CASTLING for tagging 215 nuclear proteins with a green fluorescent protein. ( a ) Three oligonucleotide pools of the same design (1,577 sequences, Supplementary Table 1 ) were used to create four tag libraries by CASTLING in duplicate sampling the indicated amount of starting material for PCR. ( b ) Detected oligonucleotide sequences of the design after PCR amplification (blue), self-integrating cassette (SIC) assembly (green) and in the final library (orange); oligonucleotides with copy number estimates (unique UMI counts) in the lowest quartile (lower 25%) are shown in light shade. ( c ) Same as (b), but evaluated in terms of open reading frames (ORFs) represented by the oligonucleotides or SICs. ( d ) Copy number of PCR amplicons recovered (red) or lost (blue) after recombineering; black horizontal lines indicate median UMI counts. ( e ) Pearson’s pairwise correlation of oligonucleotide or SIC copy number between replicates after PCR or rolling-circle amplification (RCA) respectively; n.s., not significant (p > 0.05). ( f ) Kernel density estimates of copy number in replicate 1a as normalized to the median copy number observed in the oligonucleotide pool (before recombineering) and after recombineering into the SIC pool (left panel); the distribution of fold-changes (right panel) highlights two frequency ranges: [0.1–0.9], i.e. 80% of SICs, and [0.25–0.75], i.e. 50% of SICs. ( g ) Representative fluorescence microscopy images of cells displaying nuclear, diffuse non-nuclear (asterisks), or no mNeonGreen fluorescence (arrows); scale bar 5 µm. ( h ) Quantification of fluorescence localization in > 1,000 cells in each replicate. ( i ) Recurrence of off-target events as revealed by Anchor-Seq across all library replicates and all genomic loci (left panel); the fraction of cells with SICs integrated at off-target sites (blue) within each clone population (red) is shown (right panel, axis trimmed). Panels b–i: Source data are provided as a Source Data file.

    Article Snippet: This would exceed available genome-wide tagging collection, e.g. the C-GFP collection with 4,159 ORFs (Thermo Fisher), the TAP-tag collection with 4,247 ORFs (Dharmacon) or our tandem fluorescent timer collection with 4,081 ORFs .

    Techniques: Sampling, Polymerase Chain Reaction, Amplification, Fluorescence, Microscopy

    CRISPR-Cas12a-assisted single gene-tagging in yeast. ( a ) After transformation of the self-integrating cassette (SIC) into a cell, the crRNA expressed from the SIC directs a CRISPR-Cas12a endonuclease to the genomic target locus where the DNA double-strand is cleaved. The lesion is repaired by homologous recombination using the SIC as repair template so that an in-frame gene fusion is observed. ( b ) Efficiency of seven SICs of C-terminal tagging of highly expressed open-reading frames (ORFs) with a fluorescent protein reporter, in the absence (grey) or presence (purple) of FnCas12a. Colony-forming units (CFUs) per microgram of DNA and cells used for transformation, and integration fidelity by colony fluorescence are shown. ( c ) Co-integration events upon simultaneous transformation of two SICs directed against either ENO2 or PDC1 . Both SICs confer resistance to Geneticin (G-418), but contain different fluorescent protein tags. Colonies exhibiting green and red fluorescence (arrows) were streaked to identify true co-integrands. False-color fluorescence microscopy images show nuclear Pdc1-GFP in green and the cytosolic Eno2-RFP in magenta; scale bar 5 µm. ( d ) Titration of both SICs against each other (lower panel) with evaluation of GFP-tagged (GFP+), RFP-tagged (RFP+) or co-transformed (GFP+RFP+) colonies. Panels b–d: Source data are provided as a Source Data file.

    Journal: bioRxiv

    Article Title: Pooled clone collections by multiplexed CRISPR-Cas12a-assisted gene tagging in yeast

    doi: 10.1101/476804

    Figure Lengend Snippet: CRISPR-Cas12a-assisted single gene-tagging in yeast. ( a ) After transformation of the self-integrating cassette (SIC) into a cell, the crRNA expressed from the SIC directs a CRISPR-Cas12a endonuclease to the genomic target locus where the DNA double-strand is cleaved. The lesion is repaired by homologous recombination using the SIC as repair template so that an in-frame gene fusion is observed. ( b ) Efficiency of seven SICs of C-terminal tagging of highly expressed open-reading frames (ORFs) with a fluorescent protein reporter, in the absence (grey) or presence (purple) of FnCas12a. Colony-forming units (CFUs) per microgram of DNA and cells used for transformation, and integration fidelity by colony fluorescence are shown. ( c ) Co-integration events upon simultaneous transformation of two SICs directed against either ENO2 or PDC1 . Both SICs confer resistance to Geneticin (G-418), but contain different fluorescent protein tags. Colonies exhibiting green and red fluorescence (arrows) were streaked to identify true co-integrands. False-color fluorescence microscopy images show nuclear Pdc1-GFP in green and the cytosolic Eno2-RFP in magenta; scale bar 5 µm. ( d ) Titration of both SICs against each other (lower panel) with evaluation of GFP-tagged (GFP+), RFP-tagged (RFP+) or co-transformed (GFP+RFP+) colonies. Panels b–d: Source data are provided as a Source Data file.

    Article Snippet: This would exceed available genome-wide tagging collection, e.g. the C-GFP collection with 4,159 ORFs (Thermo Fisher), the TAP-tag collection with 4,247 ORFs (Dharmacon) or our tandem fluorescent timer collection with 4,081 ORFs .

    Techniques: CRISPR, Transformation Assay, Homologous Recombination, Fluorescence, Microscopy, Titration

    Rsu-1 levels in HCC. A) Graphical representation of the number of HCC cases positive, negative or moderately positive for Rsu-1 in HCC tissue array (24 cases/48 cores). B) Protein levels of Rsu-1 in HCC cell lines compared to human hepatocytes (HH). Most of the HCC cell lines show decrease in Rsu-1 protein. C) Successful overexpression of Rsu-GFP (fusion protein) in Hep3B cell line. GFP tagged ORF clone of Homo sapiens Rsu-1 (#RG203334, Origene) was transfected into Hep3B cell line and analyzed for Rsu-1 48 h after transfection. Since it is a GFP fused protein, the MW of Rsu-1 is ~fifty-five kd instead of twenty-nine kd (MW of GFP is ~twenty-six kd). D) GFP-Rsu-1 fusion protein associates with PINCH inside the cell. Overexpression of GFP-Rsu-1 in Hep3B cell line leads to association of GFP-Rsu-1 with PINCH. GFP was immunoprecipitated 48 h after transfection. GFP precipitates were probed with either GFP or PINCH. Presence of PINCH in GFP precipitates shows association of GFP-Rsu-1with PINCH.

    Journal: PLoS ONE

    Article Title: Role of PINCH and Its Partner Tumor Suppressor Rsu-1 in Regulating Liver Size and Tumorigenesis

    doi: 10.1371/journal.pone.0074625

    Figure Lengend Snippet: Rsu-1 levels in HCC. A) Graphical representation of the number of HCC cases positive, negative or moderately positive for Rsu-1 in HCC tissue array (24 cases/48 cores). B) Protein levels of Rsu-1 in HCC cell lines compared to human hepatocytes (HH). Most of the HCC cell lines show decrease in Rsu-1 protein. C) Successful overexpression of Rsu-GFP (fusion protein) in Hep3B cell line. GFP tagged ORF clone of Homo sapiens Rsu-1 (#RG203334, Origene) was transfected into Hep3B cell line and analyzed for Rsu-1 48 h after transfection. Since it is a GFP fused protein, the MW of Rsu-1 is ~fifty-five kd instead of twenty-nine kd (MW of GFP is ~twenty-six kd). D) GFP-Rsu-1 fusion protein associates with PINCH inside the cell. Overexpression of GFP-Rsu-1 in Hep3B cell line leads to association of GFP-Rsu-1 with PINCH. GFP was immunoprecipitated 48 h after transfection. GFP precipitates were probed with either GFP or PINCH. Presence of PINCH in GFP precipitates shows association of GFP-Rsu-1with PINCH.

    Article Snippet: GFP tagged ORF clone of human Rsu-1 (#RG203334, Origene) was used for overexpressing Rsu-1 in Hep3B cell line and analyzed for Rsu-1 48 h after transfection.

    Techniques: Over Expression, Transfection, Immunoprecipitation

    Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, HUVEC were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and VEGF dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).

    Journal: International Journal of Nanomedicine

    Article Title: Effects of Zinc Oxide Nanoparticles in HUVEC: Cyto- and Genotoxicity and Functional Impairment After Long-Term and Repetitive Exposure in vitro

    doi: 10.2147/IJN.S246797

    Figure Lengend Snippet: Experimental design of long-term exposure to ZnO NPs. Twenty-four hours after seeding, HUVEC were exposed to ZnO NPs for further 24 hrs. Then, MTT assay, comet assay and VEGF dot blot were performed ( A ). For the tube formation assay, HUVEC were seeded on Matrigel together with ZnO NPs and analyzed after 5 hrs ( B ). Analysis for the proliferation assay was performed at seeding, before treatment, after long-term treatment, 24 hrs after treatment and 48 hrs after treatment ( C ).

    Article Snippet: Functional Impairment: VEGF Dot Blot To examine the impact of ZnO NPs on the capability of HUVEC to produce and secrete cytokines such as Vascular Endothelial Growth Factor (VEGF) in the paracrine way, a dot blot assay (Raybiotec Inc., Norcross, GA, USA) was performed.

    Techniques: MTT Assay, Single Cell Gel Electrophoresis, Dot Blot, Tube Formation Assay, Proliferation Assay

    Dot blot. Dot blot indicated less VEGF release in HUVEC treated with ZnO NPs. Pictures of the negative control ( A ) and 5 µg/mL ( B ) show VEGF dots highlighted by red ellipses. Graph ( C ) shows the integrated density of the VEGF dots released by HUVEC after exposure to ECGM (c; negative control) and ZnO NPs (µg/mL) analyzed with ImageJ.

    Journal: International Journal of Nanomedicine

    Article Title: Effects of Zinc Oxide Nanoparticles in HUVEC: Cyto- and Genotoxicity and Functional Impairment After Long-Term and Repetitive Exposure in vitro

    doi: 10.2147/IJN.S246797

    Figure Lengend Snippet: Dot blot. Dot blot indicated less VEGF release in HUVEC treated with ZnO NPs. Pictures of the negative control ( A ) and 5 µg/mL ( B ) show VEGF dots highlighted by red ellipses. Graph ( C ) shows the integrated density of the VEGF dots released by HUVEC after exposure to ECGM (c; negative control) and ZnO NPs (µg/mL) analyzed with ImageJ.

    Article Snippet: Functional Impairment: VEGF Dot Blot To examine the impact of ZnO NPs on the capability of HUVEC to produce and secrete cytokines such as Vascular Endothelial Growth Factor (VEGF) in the paracrine way, a dot blot assay (Raybiotec Inc., Norcross, GA, USA) was performed.

    Techniques: Dot Blot, Negative Control

    Experimental validation of target gene predictions. Validation of putative cell cycle relevant target genes for microRNAs miR-17 ( A ), miR-20a ( B ), and miR-106b ( C ) in HEK293T-cells. 3′-UTR fragments from putative targets CCND1, CCND2, E2F1, E2F3, CDKN1A, MAPK9, PTEN, RB1, RBL1, RBL2, and WEE1 were cloned at the 3′-end of the Firefly ORF in Firefly/Renilla dual reporter vector pmirGLO. To test the influence of endogenous microRNAs, pmirGLO and pmirGLO/3′-UTR were each transfected into HEK293T-cells. Normalized Firefly -activities were compared to those of pairwise co-transfections of these vectors with the microRNA mimic of interest (miR-17, miR-20a, miR-106b, also including an unspecific mimic negative control) to test for (i) unspecific effects of the given microRNA-mimic on Firefly/Renilla per se , (ii) effects of endogenous HEK293T microRNAs (iii) for validation of the particular target prediction. Dark grey columns show normalized Firefly activities from pmirGLO/3′-UTR transfections and pmirGLO + mimics co-transfections, light grey columns those from pmirGLO/3′-UTR + mimics co-transfections. Percent reductions of Firefly activities from co-transfections of pmirGLO/3′-UTR + mimic compared to pmirGLO + mimic are given as mean values from 2 biological experiments, each consisting of 4 technical replicates. Error bars represent standard deviations and statistical significancies (Student's t -test, unpaired, ***: p≤0.001, **: p≤0.01) are given. Effects of the unspecific negative control on pmirGLO and pmirGLO/3′-UTR vectors are shown separately in Fig.S2 . Except for E2F3 and MAPK9, all predicted proteins could be validated as statistically significant targets for all three microRNAs tested, although microRNA influence varied from strong (i.e. RBL2) to moderate (i.e. WEE1).

    Journal: PLoS ONE

    Article Title: MicroRNAs MiR-17, MiR-20a, and MiR-106b Act in Concert to Modulate E2F Activity on Cell Cycle Arrest during Neuronal Lineage Differentiation of USSC

    doi: 10.1371/journal.pone.0016138

    Figure Lengend Snippet: Experimental validation of target gene predictions. Validation of putative cell cycle relevant target genes for microRNAs miR-17 ( A ), miR-20a ( B ), and miR-106b ( C ) in HEK293T-cells. 3′-UTR fragments from putative targets CCND1, CCND2, E2F1, E2F3, CDKN1A, MAPK9, PTEN, RB1, RBL1, RBL2, and WEE1 were cloned at the 3′-end of the Firefly ORF in Firefly/Renilla dual reporter vector pmirGLO. To test the influence of endogenous microRNAs, pmirGLO and pmirGLO/3′-UTR were each transfected into HEK293T-cells. Normalized Firefly -activities were compared to those of pairwise co-transfections of these vectors with the microRNA mimic of interest (miR-17, miR-20a, miR-106b, also including an unspecific mimic negative control) to test for (i) unspecific effects of the given microRNA-mimic on Firefly/Renilla per se , (ii) effects of endogenous HEK293T microRNAs (iii) for validation of the particular target prediction. Dark grey columns show normalized Firefly activities from pmirGLO/3′-UTR transfections and pmirGLO + mimics co-transfections, light grey columns those from pmirGLO/3′-UTR + mimics co-transfections. Percent reductions of Firefly activities from co-transfections of pmirGLO/3′-UTR + mimic compared to pmirGLO + mimic are given as mean values from 2 biological experiments, each consisting of 4 technical replicates. Error bars represent standard deviations and statistical significancies (Student's t -test, unpaired, ***: p≤0.001, **: p≤0.01) are given. Effects of the unspecific negative control on pmirGLO and pmirGLO/3′-UTR vectors are shown separately in Fig.S2 . Except for E2F3 and MAPK9, all predicted proteins could be validated as statistically significant targets for all three microRNAs tested, although microRNA influence varied from strong (i.e. RBL2) to moderate (i.e. WEE1).

    Article Snippet: Experimental validation of target gene predictions PCR-products of full length 3′-UTRs or fragments of 3′-UTRs covering the predicted microRNA binding sites on the target gene mRNA of interest were cloned at the 3′-end of Firefly luciferase ORF in dual reporter (Firefly and Renilla luciferases) vector pmirGLO (GenBank accession FJ376737, Promega, Mannheim, FRG) using restriction enzyme pairs SacI/XbaI or SalI/XhoI.

    Techniques: Clone Assay, Plasmid Preparation, Transfection, Negative Control

    Schematic overview of the high-throughput screening (HTS) for the determination of the bacterial interactive metabolites using the  E. coli  K-12 Keio collection.

    Journal: Scientific Reports

    Article Title: Genome-wide high-throughput screening of interactive bacterial metabolite in the algal population using Escherichia coli K-12 Keio collection

    doi: 10.1038/s41598-020-67322-w

    Figure Lengend Snippet: Schematic overview of the high-throughput screening (HTS) for the determination of the bacterial interactive metabolites using the E. coli K-12 Keio collection.

    Article Snippet: To perform the high-throughput screening of bacterial interactive metabolites, our previous study, we had used a complete set of E. coli K-12 ORF archive (ASKA) library, which is a collection of gene over-expressing bacterial cells .

    Techniques: High Throughput Screening Assay

    Flow chart of ORF prediction process whereby the translated Texas synganglion transcriptome ORFs are routed through TMHMM and GPCRPred to identify candidate GPCR-encoding sequences. The transcriptome sequences were translated into all 6 possible ORFs

    Journal: Ticks and tick-borne diseases

    Article Title: Prediction of G protein-coupled receptor encoding sequences from the synganglion transcriptome of the cattle tick, Rhipicephalus microplus

    doi: 10.1016/j.ttbdis.2016.02.014

    Figure Lengend Snippet: Flow chart of ORF prediction process whereby the translated Texas synganglion transcriptome ORFs are routed through TMHMM and GPCRPred to identify candidate GPCR-encoding sequences. The transcriptome sequences were translated into all 6 possible ORFs

    Article Snippet: The unigenes from the Texas synganglion transcriptome were further analyzed ( ) with a custom open reading frame (ORF)-finding script that examined all 6 ORFs of each unigene and subsequently output any resulting protein coding sequence having a length ≥50 amino acids.

    Techniques: Flow Cytometry