Journal: International Journal of Oncology

Article Title: EPB41L3 is a potential tumor suppressor gene and prognostic indicator in esophageal squamous cell carcinoma

doi: 10.3892/ijo.2018.4316

Figure Lengend Snippet: Reduced expression of EPB41L3 in ESCC is due to methylation. (A) Methylation-specific polymerase chain reaction analysis of the seven cell lines revealed that EPB41L3 was partially methylated in all six ESCC cell lines while it was unmethylated in the normal Het-1a cells. (B) Representation schematic of the pyrosequencing results for the methylation status of the CpG promoter in the seven cell lines tested. Gray boxes indicate exons. Vertical bars indicate CpG sites examined for methylation. Black, gray, and white circles represent hypermethylation, partial methylation and nonmethylation, respectively. The schematic illustrates the representative results of pyrosequencing of a cytosine residue(s) at −141551, −141494, −141439 and −141379 bp of the EPB41L3 promoter. (C) Treatment with 5-Aza-2-deoxygcitidine restored the expression of EPB41L3 in ESCC lines, suggesting that downregulation of EPB41L3 in ESCC is due to methylation. (D) Reverse transcription-quantitative polymerase chain reaction analyses revealed that EPB41L3 expression was increased in ESCC cell lines following 5-Aza-2-deoxygcitidine treatment. The increase was most evident in the KYSE-510 cells. In the other cell lines, although an increase was observed, this was not as evident as in the KYSE-510 cells. (E) Western blot analyses revealed that EPB41L3 expression was increased in KYSE-150 and KYSE-510 cells following 5-Aza-2-deoxygcitidine treatment. EPB41L3, erythrocyte membrane protein band 4.1 like 3; ESCC, esophageal squamous cell carcinoma; M, methylated primer; U, unmethylated primer; 5aza, 5-Aza-2-deoxygcitidine.

Article Snippet: Reverse transcription-polymerase chain reaction (RT-PCR) and reverse transcription -quantitative PCR (RT-qPCR) Using the total RNA extracted from cells described above, reverse transcription was performed using the PrimeScript RT Reagent kit with gDNA Eraser (Takara Bio Inc., Otsu, Japan). qPCR was performed using SYBR Premix Ex Taq reagent (Takara Bio Inc.).

Techniques: Expressing, Methylation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Western Blot

Journal: Oncology Letters

Article Title: Serum immunoreactivity of cancer/testis antigen OY-TES-1 and its tissues expression in glioma

doi: 10.3892/ol.2017.5799

Figure Lengend Snippet: Expression of OY-TES-1 mRNA in glioma tissue. (A) Representative 12 RT-PCR results of OY-TES-1 expression in glioma (lanes 1–12), positive control testis samples (lane P) and negative controls with no cDNA (lane N). p53 was amplified as the internal control. (B) RT-qPCR analysis of OY-TES-1 mRNA. OY-TES-1 mRNA was elevated in glioma tissues compared with normal adult tissues, with the exception of testis tissue. The levels of OY-TES-1 mRNA in glioma tissues were significantly higher than in normal brain tissues (P=0.0015). (C) Comparison of OY-TES-1 mRNA between WHO grade I–II and III–IV glioma tissues, as examined by RT-qPCR. The level of OY-TES-1 mRNA was significantly higher in high-grade compared with low-grade glioma samples (*P=0.0002 vs. WHO I–II). Results are presented as the mean ± standard deviation. RT-PCR, reverse transcription-polymerase chain reaction; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; p53, tumor protein 53; WHO, World Health Organization; HPRT, hypoxanthine phosphoribosyl transferase.

Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) The presence of OY-TES-1 mRNA was quantitatively detected using the iCycler iQ™ Multi-Color Real-Time PCR Detection system (Bio-Rad Laboratories, Inc.) with the following primer sequences: Sense, 5′-GCGACACCTCCCACAAGAC-3′ and antisense, 5′-GCCCACCGTACAAATCCAG-3′.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Positive Control, Amplification, Quantitative RT-PCR, Standard Deviation, Real-time Polymerase Chain Reaction

Journal: PLoS ONE

Article Title: Gene Expression of Protein-Coding and Non-Coding RNAs Related to Polyembryogenesis in the Parasitic Wasp, Copidosoma floridanum

doi: 10.1371/journal.pone.0114372

Figure Lengend Snippet: Results of RT-qPCR for protein coding genes. Gene specific primers for RT-qPCR were mentioned in Table S2 . The detailes of PCR condition for each target gene were descrived in Supporting Information S1 . (A)–(D): Bar graphs showing gene expression relative to gapdh (means; bars = S.D). MH: adult male head (n = 3), MA: adult male abdomen (n = 3), FH: adult female head (n = 3), FA; adult female abdomen (n = 3), cleavage: cleavage-stage embryo (1–8 h post-oviposition, n = 9), morula: primary morula-stage embryo (12–24 h post-oviposition, n = 12). No differences were determined to be statistically significant for any comparison of pairs at p = 0.05 (two-sample t-test or Welch's test).

Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) was performed using the same cDNA samples as templates with THUNDERBIRD SYBR qPCR Mix (TOYOBO) and gene-specific primers ( ).

Techniques: Quantitative RT-PCR, Polymerase Chain Reaction, Expressing

Journal: PLoS ONE

Article Title: Gene Expression of Protein-Coding and Non-Coding RNAs Related to Polyembryogenesis in the Parasitic Wasp, Copidosoma floridanum

doi: 10.1371/journal.pone.0114372

Figure Lengend Snippet: Results of RT-qPCR for CflncRNAs. Gene specific primers for RT-qPCR were mentioned in Table S2 . The detailes of PCR condition for each target gene were descrived in Supporting Information S1 . (A)–(C): Bar graphs showing the gene expression relative to gapdh (means, bars = S.D). MH: adult male head (n = 3), MA: adult male abdomen (n = 3), FH: adult female head (n = 3), FA; adult female abdomen (n = 3), cleavage: cleavage-stage embryo (1–8 h post-oviposition, n = 9), morula: primary morula-stage embryo (12–24 h post-oviposition, n = 12). For A and B, *indicates a statistically significant difference, p

Article Snippet: Reverse transcription quantitative PCR (RT-qPCR) was performed using the same cDNA samples as templates with THUNDERBIRD SYBR qPCR Mix (TOYOBO) and gene-specific primers ( ).

Techniques: Quantitative RT-PCR, Polymerase Chain Reaction, Expressing

Journal: Frontiers in Microbiology

Article Title: Plasmodium falciparumvar Gene Is Activated by Its Antisense Long Noncoding RNA

doi: 10.3389/fmicb.2018.03117

Figure Lengend Snippet: Schematic diagram of plasmids for expressing the var aslncRNA and 5′ end identification of PF3D7_0617400 aslncRNA. (A) Schematic diagram of pT7SE-as0617400 and pT7-as0617400. The translation initial site of ATG of PF3D7_0617400 is marked as +1 and the stop codon is marked with +8375, the ∼2.5 kb var fragment (from +6412 to +3952) was amplified and inserted into pT7SE and pT7, respectively. The wavy line is the natural var aslncRNA. T7 pro, T7 promoter; T7 ter, T7 terminator. (B) 5′ end identification of PF3D7_0617400 aslncRNA. PCR products of 5′ end RACE obtained according to the standard manuals. M: the DNA marker λ-EcoT14 I digest (TAKARA), lane1: PCR products of 5′ RACE. (C) The PF3D7_0617400 aslncRNA transcriptional start sites. The translation start site of PF3D7_0617400 is marked as +1 on the schematic diagram. Capital letters are the coding sequence of the var exonI and the lower letters are intron sequence. The transcriptional start sites of the aslncRNAs are marked with underlined numbers and arrows. The underlined letters locations are displayed relative to translation start site, and the arrows indicate the aslncRNA transcription directions. (D) RT-qPCR quantification of the NLS-T7RNP transcription in C8/pT7- as0617400 ( 1 ) and C8/pT7SE-as0617400 ( 2 ). Relative transcripts numbers are normalized to serine-tRNA ligase gene ( PF3D7_0717700 ). ∗ P

Article Snippet: Reverse Transcription Quantitative PCR (RT-qPCR) One microgram of the extracted total RNA of P. falciparum was reverse transcribed using FastQuant RT Kit (Tiangen) according to its standard manuals.

Techniques: Expressing, Amplification, Polymerase Chain Reaction, Marker, Sequencing, Quantitative RT-PCR

Journal: Applied and Environmental Microbiology

Article Title: Whole-Transcriptome Shotgun Sequencing (RNA-seq) Screen Reveals Upregulation of Cellobiose and Motility Operons of Lactobacillus ruminis L5 during Growth on Tetrasaccharides Derived from Barley ?-Glucan

doi: 10.1128/AEM.01887-13

Figure Lengend Snippet: Extended transcriptional analysis measured by reverse transcription-quantitative PCR (RT-qPCR).

Article Snippet: Reverse transcription-quantitative PCR (RT-qPCR) was carried out using 384-well plates (4titude Ltd. Surrey, United Kingdom) with Optical adhesive film (Life Technologies, Foster City, CA, USA) and an ABI Viia-7 Fast system.

Techniques: Real-time Polymerase Chain Reaction, Quantitative RT-PCR