Journal: Cell Death & Disease

Article Title: MicroRNA-221-3p, a TWIST2 target, promotes cervical cancer metastasis by directly targeting THBS2

doi: 10.1038/s41419-017-0077-5

Figure Lengend Snippet: a SiHa and HeLa cells were transfected with miR-221-3p mimic, miR-221-3p inhibitor, miR-221-3p mimic-nc, and miR-221-3p inhibitor-nc. Western blotting analysis of E-cadherin, N-cadherin, and Vimentin was performed of the five groups in SiHa and HeLa cells. β-actin was used as loading control. b qRT-PCR analysis of E-cadherin, N-cadherin, and Vimentin of the five groups in SiHa and HeLa cells. c Wound-healing assay of the five groups in SiHa and HeLa cells. d Boyden chamber assay of the five groups in SiHa and HeLa cells. e Cell migration was quantified as percentage of wound-healed area. f Average number of invading cells per field from three independent experiments. Data represent means ± SD of five randomly selected areas. * p < 0.05

Article Snippet: Human CC cell lines SiHa and HeLa, and human embryonic kidney 293T cells were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Transfection, Western Blot, Control, Quantitative RT-PCR, Wound Healing Assay, Boyden Chamber Assay, Migration

Journal: Cell Death & Disease

Article Title: MicroRNA-221-3p, a TWIST2 target, promotes cervical cancer metastasis by directly targeting THBS2

doi: 10.1038/s41419-017-0077-5

Figure Lengend Snippet: a Correlation of bioluminescence intensity with cell quantity in SiHa-luc-RFP-221-3p, SiHa-luc-RFP-nc, HeLa-luc-RFP-221-3p, and HeLa-luc-RFP-NC cells in vitro . b In vivo bioluminescence images of lymphatic metastasis. Primary tumour (P) in claw pad of mice and metastases (M). Lymph nodes in popliteal and inguinal regions were measured by detecting bioluminescence signals. The ratio of metastasis-positive lymph nodes was calculated. * p < 0.05. c In situ hybridization of miR-221-3p expression in primary tumors of mice injected with SiHa-luc-RFP-221-3p, SiHa-luc-RFP-nc, HeLa-luc-RFP-221-3p or HeLa-luc-RFP-NC cells. d Primary tumour site and lymph nodes with tumour cells were identified by tumour cell-expressed RFP

Article Snippet: Human CC cell lines SiHa and HeLa, and human embryonic kidney 293T cells were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: In Vitro, In Vivo, In Situ Hybridization, Expressing, Injection

Journal: Cell Death & Disease

Article Title: MicroRNA-221-3p, a TWIST2 target, promotes cervical cancer metastasis by directly targeting THBS2

doi: 10.1038/s41419-017-0077-5

Figure Lengend Snippet: a Bioinformatic prediction and screening of potential transcription factors of miR-221-3p. b Correlation analysis of TWIST2 and miR-221-3p expression detected by qRT-PCR in 28 additional SCC tissues. The expression levels of TWIST2 were plotted against the expression levels of miR-221-3p. c The certified result of microarray analysis. Hierarchical clustering of eight significantly dysregulated miRNAs expression profiles of the three groups, including SiHa, SiHa-shtw2, and SiHa-tw2 cells. d qRT-PCR analysis of the eight significantly dysregulated miRNAs expressed in SiHa, SiHa-shtw2, and SiHa-tw2 cells, and in HeLa cells transfected with TWIST2 expression vector or TWIST2 siRNA. MiR-221-3p emerged as a highly upregulated miRNA. Data represent means ± SD of five randomly selected areas (* p < 0.05)

Article Snippet: Human CC cell lines SiHa and HeLa, and human embryonic kidney 293T cells were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Expressing, Quantitative RT-PCR, Microarray, Transfection, Plasmid Preparation

Journal: Cell Death & Disease

Article Title: MicroRNA-221-3p, a TWIST2 target, promotes cervical cancer metastasis by directly targeting THBS2

doi: 10.1038/s41419-017-0077-5

Figure Lengend Snippet: a Schematic structure of the miR-221-3p upstream promoter containing a TWIST2-binding site. b Luciferase activity of PGL3-221-3p construct after transfection of TWIST2 plasmid in SiHa and 293 T cells. c Western blotting analysis of expression of E-cadherin, N-cadherin, and Vimentin proteins in SiHa, SiHa-tw2, SiHa-shwt2, or SiHa-shtw2 cells transfected with miR-221-3p mimic or miR-221-3p inhibitor; and HeLa cells transfected with TWIST2 expression vector with/without miR-221-3p mimic or TWIST2 siRNA with/without miR-221-3p inhibitor. β-actin was used as loading control. d Cell migration was quantified as the percentage of wound-healed area; average number of invaded cells per field from three independent experiments. Data represent means ± SD of five randomly selected areas (* p < 0.05; ** p < 0.001). e IHC analysis of TWIST2 and in situ hybridization analysis of miR-221-3p in matched cervical specimens. Human CC tumors with high TWIST2 expression and had strong miR-221-3p expression. Normal cervical tissues with low TWIST2 expression and had weak miR-221-3p expression. Representative images are shown at ×400 magnification. The HSCOREs of miR-221-3p and TWIST2 in different cervical specimens are also shown (* p < 0.05) (upper-right panel). Correlation of miR-221-3p staining and TWIST2 staining was analyzed (lower-right panel)

Article Snippet: Human CC cell lines SiHa and HeLa, and human embryonic kidney 293T cells were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Binding Assay, Luciferase, Activity Assay, Construct, Transfection, Plasmid Preparation, Western Blot, Expressing, Control, Migration, In Situ Hybridization, Staining

Journal: Cell Death & Disease

Article Title: MicroRNA-221-3p, a TWIST2 target, promotes cervical cancer metastasis by directly targeting THBS2

doi: 10.1038/s41419-017-0077-5

Figure Lengend Snippet: a Bioinformatic prediction and screening of potential miRNAs targeting miR-221-3p; incomplete complementation of the base of miR-221-3p to the 3′UTR region of THBS2 mRNA. Also shown are nucleotides mutated in THBS2 mutant 3′UTR. b Luciferase activity of wild-3′UTR-THBS2-luc and mutant 3′UTR-THBS2-luc constructs in SiHa or 293 T cells after transfection of miR-221-3p mimic. c Western blotting analysis of the expression of THBS2 in SiHa and HeLa cells transfected with miR-221-3p mimic, miR-221-3p inhibitor, miR-221-3p mimic-nc, and miR-221-3p inhibitor-nc. β-actin was used as loading control. d The HSCOREs of THBS2 in different cervical specimens (left), and correlation of THBS2 staining and miR-221-3p was analyzed (right). e Normal cervical tissues with high THBS2 expression. Human CC tumors with low THBS2 expression. * p < 0.05

Article Snippet: Human CC cell lines SiHa and HeLa, and human embryonic kidney 293T cells were purchased from American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Mutagenesis, Luciferase, Activity Assay, Construct, Transfection, Western Blot, Expressing, Control, Staining

Journal: Microbiology

Article Title: Global transcriptional analysis of the stringent response in Enterococcus faecalis

doi: 10.1099/mic.0.060236-0

Figure Lengend Snippet: qRT-PCR primers used to validate the microarray results

Article Snippet: The slides were hybridized to a mixture containing equal amounts of test and reference cDNA for 16 h at 42 °C in a MicroArray User Interface (MAUI) hybridization chamber (BioMicro Systems).

Techniques: Microarray

Journal: Microbiology

Article Title: Global transcriptional analysis of the stringent response in Enterococcus faecalis

doi: 10.1099/mic.0.060236-0

Figure Lengend Snippet: qRT-PCR validation of microarray data

Article Snippet: The slides were hybridized to a mixture containing equal amounts of test and reference cDNA for 16 h at 42 °C in a MicroArray User Interface (MAUI) hybridization chamber (BioMicro Systems).

Techniques: Biomarker Discovery, Microarray

Journal: BMC Microbiology

Article Title: Separation of the bacterial species, Escherichia coli , from mixed-species microbial communities for transcriptome analysis

doi: 10.1186/1471-2180-11-59

Figure Lengend Snippet: Plot of gene expression of sorted/unsorted cells . Plot of one-sample T-test p-values with fold-change in gene expression for all ORFs in microarray study I. Vertical lines show the cutoff of fold-change of 2 (Log 2 ratio of ± 1), while the horizontal line shows the cutoff of p-value 0.05. Genes located in the left-bottom corner (Log 2 ratio <-1 and p-value <0.05) and in the right-bottom corner (Log 2 ratio >1 and p-value <0.05) were considered to have their expressions changed due to dispersion/homogenization and IMS (immuno-magnetic separation) cell sorting. A total of ten genes were selected using these criteria, eight of which also differentially expressed in the independent microarray study II.

Article Snippet: Hybridization was in a Corning Microarray Hybridization Chamber (Corning Inc.) in 42°C water bath.

Techniques: Expressing, Microarray, Homogenization, FACS

Journal: BMC Microbiology

Article Title: Separation of the bacterial species, Escherichia coli , from mixed-species microbial communities for transcriptome analysis

doi: 10.1186/1471-2180-11-59

Figure Lengend Snippet: Genes identified as differentially expressed # between IMS sorted E. coli cells versus unsorted E. coli cells* by the method of cDNA microarray and their differential expression confirmed with another method of qPCR

Article Snippet: Hybridization was in a Corning Microarray Hybridization Chamber (Corning Inc.) in 42°C water bath.

Techniques: Microarray, Expressing

Journal:

Article Title: A microarray-based high throughput molecular marker genotyping method: the tagged microarray marker (TAM) approach

doi: 10.1093/nar/gng113

Figure Lengend Snippet: The tagged microarray approach. (a) Two alleles are distinguished by PCR using three primers. The biotinylated PCR products are arrayed onto a solid support and hybridised with detector probes, which recognise tags specific to the two products. (b) Structure of a tag primer. The allele-specific region is separated from the tag by a C18 linker. (c) Tag detector probe. Two partially complementary oligonucleotides carry a tag sequence (the A tag in this case), its reverse complement (A′), a region of cross-homology (B and B′ for this pair) and a fluorochrome each.

Article Snippet: Slides were hybridised in a Grant microarray hybridisation chamber (In Slide Out hybridisation oven; Boekel Scientific, Feasterville, PA) at 100% relative humidity at 37°C for 30 min.

Techniques: Microarray, Sequencing

Journal:

Article Title: A microarray-based high throughput molecular marker genotyping method: the tagged microarray marker (TAM) approach

doi: 10.1093/nar/gng113

Figure Lengend Snippet: The tagged microarray approach applied to a pea genomic retrotransposon insertion. (a) The two alleles differ by the presence or absence of a retrotransposon insertion (PDR1 here). Different PCR products are produced from the two alleles and these can be recognised either by gel electrophoresis or the tagged microarray approach (Fig. ​(Fig.1).1). (b) Microarray image from 384 pea samples assayed for the 1794-1 allele. Each sample was spotted four times in a row. Cy3 fluorescing spots (green) represent the occupied site allele and Cy5 labelled spots (red) show the unoccupied site allele. Arrowed samples were analysed by gel electrophoresis (Fig. ​(Fig.1c).1c). (c) Agarose gel of samples arrowed in (b), together with corresponding fluorescent images taken from the array image. (d) Quantitation of array spot intensities shown in (c). SD, standard deviation.

Article Snippet: Slides were hybridised in a Grant microarray hybridisation chamber (In Slide Out hybridisation oven; Boekel Scientific, Feasterville, PA) at 100% relative humidity at 37°C for 30 min.

Techniques: Microarray, Produced, Nucleic Acid Electrophoresis, Agarose Gel Electrophoresis, Quantitation Assay, Standard Deviation

Journal:

Article Title: A microarray-based high throughput molecular marker genotyping method: the tagged microarray marker (TAM) approach

doi: 10.1093/nar/gng113

Figure Lengend Snippet: Modifications to the tagged microarray approach. (a) Using tag detectors labelled with different fluorochromes produces different fluorescent outputs from the same samples. Sample JI 2698 contains an occupied 1794-1 allele and sample Therèse is unoccupied. Samples were spotted in triplicate. (b) Using four tag detectors to analyse the allelic state for two loci in the same reaction. Sample JI 2698 is occupied for 1794-1 and unoccupied for UniTpv, sample Therèse is unoccupied for 1794-1 and occupied for UniTpv and sample JI 2055 is unoccupied for both loci. Tag detectors used for both loci produce Cy3 signal (green) from an occupied allele and Cy5 signal (red) from an unoccupied allele.

Article Snippet: Slides were hybridised in a Grant microarray hybridisation chamber (In Slide Out hybridisation oven; Boekel Scientific, Feasterville, PA) at 100% relative humidity at 37°C for 30 min.

Techniques: Microarray

Journal:

Article Title: A microarray-based high throughput molecular marker genotyping method: the tagged microarray marker (TAM) approach

doi: 10.1093/nar/gng113

Figure Lengend Snippet: Microarray image for 1536 pea DNA samples assayed for the 1794-1 allele under the same conditions as Figure ​Figure2b.2b. Red (Cy5) spots indicate samples containing an unoccupied (–) allele, green indicates occupied (+) and yellow indicates both alleles (+/–). Each PCR was spotted once per array.

Article Snippet: Slides were hybridised in a Grant microarray hybridisation chamber (In Slide Out hybridisation oven; Boekel Scientific, Feasterville, PA) at 100% relative humidity at 37°C for 30 min.

Techniques: Microarray

Journal:

Article Title: A microarray-based high throughput molecular marker genotyping method: the tagged microarray marker (TAM) approach

doi: 10.1093/nar/gng113

Figure Lengend Snippet: Detection of barley SNP polymorphisms using the tagged microarray approach. Cultivars Steptoe and Morex have contrasting alleles for SNP 206 and SNP 957. Cy5 fluorescence indicates the presence of one allele and Cy3 the other. Samples were spotted in triplicate. The Cy3 (green) and Cy5 (red) colour-separated images and the two-colour (overlay) image are shown. Quantification of averaged array intensities (in arbitrary fluorescence units) and the Cy3/Cy5 ratios of signal intensities are also shown. SD, standard deviation.

Article Snippet: Slides were hybridised in a Grant microarray hybridisation chamber (In Slide Out hybridisation oven; Boekel Scientific, Feasterville, PA) at 100% relative humidity at 37°C for 30 min.

Techniques: Microarray, Fluorescence, Standard Deviation

Journal:

Article Title: A microarray-based high throughput molecular marker genotyping method: the tagged microarray marker (TAM) approach

doi: 10.1093/nar/gng113

Figure Lengend Snippet: Microarray image for 384 barley DNA samples assayed for the SNP 206 allele. Reaction and detection conditions were the same as for Figure ​Figure55 and arraying format is the same as for Figure ​Figure2b.2b. Each PCR was spotted four times per array.

Article Snippet: Slides were hybridised in a Grant microarray hybridisation chamber (In Slide Out hybridisation oven; Boekel Scientific, Feasterville, PA) at 100% relative humidity at 37°C for 30 min.

Techniques: Microarray