Structured Review

Thermo Fisher oligo dt
Olfactory cng channel mRNAs are expressed in the brain. Semiquantitative RT-PCR assays were used to examine expression of mRNAs encoding rOCNC1 ( A ), rOCNC2 ( B ), and rRCNC1 ( C ). Primers specific for the 3′ untranslated regions of the three mRNAs were used to amplify <t>oligo-dT-primed</t> <t>cDNA</t> made from RNA from various adult rat tissues (35 cycles of amplification). In D , primers spanning an intron/exon boundary from the rOCNC2 gene were used for amplification, to control for the possible presence of genomic DNA in the mRNA preparations. The cDNA concentrations were normalized by amplification (15 and 20 cycles) with β-actin primers, and ∼1 ng of cDNA was used per amplification. Lanes: M , DNA markers; 1–9 , cDNAs from 1 , cortex; 2 , liver; 3 , olfactory bulb; 4 , nasal epithelium; 5 , brainstem; 6 , cerebellum; 7 , hippocampus; 8 , eye; 9 , heart; 10 , genomic rat DNA; 11 , no DNA. The positions of the appropriate amplified bands are indicated by arrows . The rOCNC1 PCR product is 159 bp ( A ); the rOCNC2 product is 122 bp ( B ); the rRCNC1 product is 202 bp ( C ); and the genomic intron product is ∼363 bp ( D ). The figure is a negative image of an ethidium bromide-stained agarose gel.
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1) Product Images from "Functional Expression of the Heteromeric “Olfactory” Cyclic Nucleotide-Gated Channel in the Hippocampus: A Potential Effector of Synaptic Plasticity in Brain Neurons"

Article Title: Functional Expression of the Heteromeric “Olfactory” Cyclic Nucleotide-Gated Channel in the Hippocampus: A Potential Effector of Synaptic Plasticity in Brain Neurons

Journal: The Journal of Neuroscience

doi: 10.1523/JNEUROSCI.17-06-01993.1997

Olfactory cng channel mRNAs are expressed in the brain. Semiquantitative RT-PCR assays were used to examine expression of mRNAs encoding rOCNC1 ( A ), rOCNC2 ( B ), and rRCNC1 ( C ). Primers specific for the 3′ untranslated regions of the three mRNAs were used to amplify oligo-dT-primed cDNA made from RNA from various adult rat tissues (35 cycles of amplification). In D , primers spanning an intron/exon boundary from the rOCNC2 gene were used for amplification, to control for the possible presence of genomic DNA in the mRNA preparations. The cDNA concentrations were normalized by amplification (15 and 20 cycles) with β-actin primers, and ∼1 ng of cDNA was used per amplification. Lanes: M , DNA markers; 1–9 , cDNAs from 1 , cortex; 2 , liver; 3 , olfactory bulb; 4 , nasal epithelium; 5 , brainstem; 6 , cerebellum; 7 , hippocampus; 8 , eye; 9 , heart; 10 , genomic rat DNA; 11 , no DNA. The positions of the appropriate amplified bands are indicated by arrows . The rOCNC1 PCR product is 159 bp ( A ); the rOCNC2 product is 122 bp ( B ); the rRCNC1 product is 202 bp ( C ); and the genomic intron product is ∼363 bp ( D ). The figure is a negative image of an ethidium bromide-stained agarose gel.
Figure Legend Snippet: Olfactory cng channel mRNAs are expressed in the brain. Semiquantitative RT-PCR assays were used to examine expression of mRNAs encoding rOCNC1 ( A ), rOCNC2 ( B ), and rRCNC1 ( C ). Primers specific for the 3′ untranslated regions of the three mRNAs were used to amplify oligo-dT-primed cDNA made from RNA from various adult rat tissues (35 cycles of amplification). In D , primers spanning an intron/exon boundary from the rOCNC2 gene were used for amplification, to control for the possible presence of genomic DNA in the mRNA preparations. The cDNA concentrations were normalized by amplification (15 and 20 cycles) with β-actin primers, and ∼1 ng of cDNA was used per amplification. Lanes: M , DNA markers; 1–9 , cDNAs from 1 , cortex; 2 , liver; 3 , olfactory bulb; 4 , nasal epithelium; 5 , brainstem; 6 , cerebellum; 7 , hippocampus; 8 , eye; 9 , heart; 10 , genomic rat DNA; 11 , no DNA. The positions of the appropriate amplified bands are indicated by arrows . The rOCNC1 PCR product is 159 bp ( A ); the rOCNC2 product is 122 bp ( B ); the rRCNC1 product is 202 bp ( C ); and the genomic intron product is ∼363 bp ( D ). The figure is a negative image of an ethidium bromide-stained agarose gel.

Techniques Used: Reverse Transcription Polymerase Chain Reaction, Expressing, Amplification, Polymerase Chain Reaction, Staining, Agarose Gel Electrophoresis

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Random Hexamer Labeling:

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Synthesized:

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Modification:

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Derivative Assay:

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Sequencing:

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Cell Culture:

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Cytokine Assay:

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Aqueous Normal-phase Chromatography:

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Polymerase Chain Reaction:

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Article Title: Contribution of the Cytoplasmic Determinants of Vpu to the Expansion of Virus-Containing Compartments in HIV-1-Infected Macrophages
Article Snippet: For each sample, cDNA synthesis was performed using 100 ng of total RNA with oligo(dT) and a high-capacity cDNA reverse transcription kit (Applied Biosystems). .. The PCR conditions and cycles were as follows: an initial DNA denaturation step at 95°C for 10 min, 45 cycles at 95°C for 10 s, an annealing step at 59°C for 30 s, and an extension step at 72°C for 10 s, followed by a melting-curve analysis cycle.

Article Title: Synthesis and Evaluation of Esterified Hsp70 Agonists in Cellular Models of Protein Aggregation and Folding
Article Snippet: To synthesize cDNA, 0.5 μg of each sample was combined with oligo dT (Thermo Scientific: 25 ng/uL) and incubated at 80oC for 3 minutes. .. Next, 50 U of Murine Leukemia Virus reverse transcriptase (ThermoFischer), 20 Units of RNase inhibitor (Qiagen), and 2 mM deoxynucleotides were added to each sample, followed by incubation at oC for 1 hr and then at 92oC for 10 min, based on the RETROscript RT kit protocol (ThermoFisher). cDNAs were diluted 1:40 and PCR was carried out on a StepOnePlus with gene-specific primers and the SYBR green reagent using the following parameters: 10 min at 95oC, then 40 cycles of 15 min at 95oC and 1 min at 60oC.

Recombinant:

Article Title: A glycan shield on chimpanzee CD4 protects against infection by primate lentiviruses (HIV/SIV)
Article Snippet: Extracted RNA was used as a template for oligo(dT) primed reverse transcription (SuperScript III RT; Thermo Fisher). .. PCR clean-up was performed using Exonuclease-I and recombinant Shrimp Alkaline Phosphatase treatment (Affymetrix) for 15 min at 37 °C, and then the cleaned-up products were Sanger sequenced using the following sequencing primers: CD4-forward 5′-GGGCAAGAAAGACGCAAGC-3′, 5′-TTCTGCGGGCTCAGGTCC-3′, 5′-TAGTGTTCGGATTGACTGC-3′, 5′-TACCCAGGACCCTAAGC-3′, and CD4-reverse 5′-ATCTGCCTGGCCTCGTG-3′, 5′-TTTACCCCTTGGACTCC-3′, 5′-GGTTCACTTCCTGATGC-3′.

Isolation:

Article Title: Inhibiting pyrimidine biosynthesis impairs Ebola virus replication through depletion of nucleoside pools and activation of innate immune responses
Article Snippet: The isolated RNA was also subjected to DNase treatment using Ambion DNA-free DNase Treatment and Removal Reagents. .. One microgram of total RNA was subjected to cDNA synthesis using oligo(dT) using the SuperScript III first strand cDNA Synthesis Kit following manufacturer’s recommendations (Life Technologies).

Article Title: Metformin inhibits pancreatic cancer cell and tumor growth and downregulates Sp transcription factors
Article Snippet: Total RNA was isolated from Panc1 cells using the RNeasy Protect Mini kit (Qiagen, Valencia, CA) according to the manufacturer’s protocol. .. Complementary DNA was prepared using a combination of oligodeoxythymidylic acid (Applied Biosystems, Foster City, CA), deoxynucleoside triphosphate mix and Superscript II reverse transcriptase (Invitrogen).

Article Title: DRUGS THAT TARGET SPECIFICITY PROTEINS DOWNREGULATE EPIDERMAL GROWTH FACTOR RECEPTOR IN BLADDER CANCER CELLS
Article Snippet: Total RNA was isolated using the RNeasy Protect Mini kit (Qiagen, Valencia, CA) according to the manufacturer’s protocol. .. RNA was reverse-transcribed using Superscript II reverse transcriptase (Invitrogen) according to the manufacturer’s protocol. cDNA was prepared from the 253JB-V and KU7 bladder cancer cell lines at different time intervals using a combination of oligodeoxythymidylic acid and dNTP mix (Applied Biosystems, Foster City, CA) and Superscript II.

Article Title: Influence of hypoxia-inducible factor 1α on dendritic cell differentiation and migration
Article Snippet: BMDCs were either stimulated or not with 1 μg/mL LPS for 3 h. RNA was isolated using the RNeasy mini kit (Qiagen, Hilden, Germany). .. First-strand cDNA was synthesized from 1 μg of total RNA using a mixture of oligo(dT) and Revert Aid reverse transcriptase (Fermentas, St.Leon-Rot, Germany).

Article Title: Pregnancy upregulates angiotensin type 2 receptor expression and increases blood flow in uterine arteries of rats
Article Snippet: Paragraph title: RNA isolation and quantitative real-time PCR ... Two micrograms of total RNA were reverse transcribed using a modified Maloney murine leukemia virus-derived reverse transcriptase (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen, Grand Island, NY).

Article Title: A Promising Therapeutic Approach for Multiple Sclerosis: Recombinant T-Cell Receptor Ligands Modulate Experimental Autoimmune Encephalomyelitis by Reducing Interleukin-17 Production and Inhibiting Migration of Encephalitogenic Cells into the CNS
Article Snippet: .. Alternatively, total RNA was isolated from sorted GFP+ cells using the RNeasy mini-kit protocol (Qiagen) and converted to cDNA using oligo(dT), random hexamers, and Superscript RT II enzyme (Invitrogen, Grand Island, NY). .. Real-time PCR for IL-17 and TNFα was performed using Quantitect SYBR Green PCR master mix (Qiagen) and primers (synthesized by Applied Biosystems, Foster City, CA).

Purification:

Article Title: Testosterone plays a permissive role in angiotensin II-induced hypertension and cardiac hypertrophy in male rats
Article Snippet: All RNA isolates were made DNA-free by treatment with DNAse and further purified with RNeasy clean up kit (QIAGEN Inc, Valencia, CA). .. One microgram of total RNA was reverse transcribed using a modified Maloney murine leukemia virus-derived RT (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen).

Article Title: Pregnancy upregulates angiotensin type 2 receptor expression and increases blood flow in uterine arteries of rats
Article Snippet: All RNA isolates were made DNA free by treatment with DNase and further purification with RNeasy Clean Up Kit (QIAGEN Inc, Valencia, CA). .. Two micrograms of total RNA were reverse transcribed using a modified Maloney murine leukemia virus-derived reverse transcriptase (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen, Grand Island, NY).

Reverse Transcription Polymerase Chain Reaction:

Article Title: Functional Expression of the Heteromeric “Olfactory” Cyclic Nucleotide-Gated Channel in the Hippocampus: A Potential Effector of Synaptic Plasticity in Brain Neurons
Article Snippet: Reverse transcription (RT)-PCR assays. .. First-strand cDNA was primed with oligo-dT from 25 μg of total RNA pretreated with DNaseI. cDNA was synthesized using the SuperScript II enzyme (Life Technologies) in 50 μl reactions at 42°C for 2 hr.

Article Title: Involvement of estradiol-17β and its membrane receptor, G protein coupled receptor 30 (GPR30) in regulation of oocyte maturation in zebrafish, Danio rerio
Article Snippet: Paragraph title: RT-PCR ... Total RNA (1–3 μg) was reverse transcribed into cDNA at 50°C for 1 h in 10 μl reaction solution containing 1 × First Strand Buffer, 10 mM dithiothreitol, 0.5 mM each dNTP, 0.5 μg oligo-dT, and 100 U SuperScript III (Invitrogen).

Article Title: Synthesis and Evaluation of Esterified Hsp70 Agonists in Cellular Models of Protein Aggregation and Folding
Article Snippet: Paragraph title: 2.7. Quantification of message levels by RT-PCR ... To synthesize cDNA, 0.5 μg of each sample was combined with oligo dT (Thermo Scientific: 25 ng/uL) and incubated at 80oC for 3 minutes.

Activated Clotting Time Assay:

Article Title: Functional Expression of the Heteromeric “Olfactory” Cyclic Nucleotide-Gated Channel in the Hippocampus: A Potential Effector of Synaptic Plasticity in Brain Neurons
Article Snippet: First-strand cDNA was primed with oligo-dT from 25 μg of total RNA pretreated with DNaseI. cDNA was synthesized using the SuperScript II enzyme (Life Technologies) in 50 μl reactions at 42°C for 2 hr. .. The rOCNC2 intron primer sequence is 5′-CAG AAG GCA AGC ACT GAA TGA (GenBank ), and this was used together with a cDNA primer (5′-GGC CAC CAG GTA ACT GTG CTG) to test for the presence of genomic DNA.

In Situ Hybridization:

Article Title: Functional Expression of the Heteromeric “Olfactory” Cyclic Nucleotide-Gated Channel in the Hippocampus: A Potential Effector of Synaptic Plasticity in Brain Neurons
Article Snippet: First-strand cDNA was primed with oligo-dT from 25 μg of total RNA pretreated with DNaseI. cDNA was synthesized using the SuperScript II enzyme (Life Technologies) in 50 μl reactions at 42°C for 2 hr. .. In situ hybridization to tissue sections.

Software:

Article Title: DRUGS THAT TARGET SPECIFICITY PROTEINS DOWNREGULATE EPIDERMAL GROWTH FACTOR RECEPTOR IN BLADDER CANCER CELLS
Article Snippet: RNA was reverse-transcribed using Superscript II reverse transcriptase (Invitrogen) according to the manufacturer’s protocol. cDNA was prepared from the 253JB-V and KU7 bladder cancer cell lines at different time intervals using a combination of oligodeoxythymidylic acid and dNTP mix (Applied Biosystems, Foster City, CA) and Superscript II. .. The ABI Dissociation Curves software was used after a brief thermal protocol (95°C for 15 s and 60°C for 20 s, followed by a slow ramp to 95°C) to control for multiple species in each PCR amplification.

Real-time Polymerase Chain Reaction:

Article Title: Testosterone plays a permissive role in angiotensin II-induced hypertension and cardiac hypertrophy in male rats
Article Snippet: One microgram of total RNA was reverse transcribed using a modified Maloney murine leukemia virus-derived RT (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen). .. One microliter of the diluted cDNA corresponding to 100 ng RNA was amplified by real-time PCR using FAM (Invitrogen) as the fluorophore in a CFX96 real-time thermal cycler (Bio-Rad, Hercules, CA).

Article Title: Inhibiting pyrimidine biosynthesis impairs Ebola virus replication through depletion of nucleoside pools and activation of innate immune responses
Article Snippet: One microgram of total RNA was subjected to cDNA synthesis using oligo(dT) using the SuperScript III first strand cDNA Synthesis Kit following manufacturer’s recommendations (Life Technologies). .. Quantitative PCR reactions were performed using SYBR green Fast (Applied Biosystems) on a Biorad Real-Time PCR machine.

Article Title: Metformin inhibits pancreatic cancer cell and tumor growth and downregulates Sp transcription factors
Article Snippet: Paragraph title: Quantitative real-time PCR and luciferase assay ... Complementary DNA was prepared using a combination of oligodeoxythymidylic acid (Applied Biosystems, Foster City, CA), deoxynucleoside triphosphate mix and Superscript II reverse transcriptase (Invitrogen).

Article Title: DRUGS THAT TARGET SPECIFICITY PROTEINS DOWNREGULATE EPIDERMAL GROWTH FACTOR RECEPTOR IN BLADDER CANCER CELLS
Article Snippet: Paragraph title: Real-Time PCR ... RNA was reverse-transcribed using Superscript II reverse transcriptase (Invitrogen) according to the manufacturer’s protocol. cDNA was prepared from the 253JB-V and KU7 bladder cancer cell lines at different time intervals using a combination of oligodeoxythymidylic acid and dNTP mix (Applied Biosystems, Foster City, CA) and Superscript II.

Article Title: Influence of hypoxia-inducible factor 1α on dendritic cell differentiation and migration
Article Snippet: Paragraph title: Quantitative real-time PCR ... First-strand cDNA was synthesized from 1 μg of total RNA using a mixture of oligo(dT) and Revert Aid reverse transcriptase (Fermentas, St.Leon-Rot, Germany).

Article Title: Pregnancy upregulates angiotensin type 2 receptor expression and increases blood flow in uterine arteries of rats
Article Snippet: Paragraph title: RNA isolation and quantitative real-time PCR ... Two micrograms of total RNA were reverse transcribed using a modified Maloney murine leukemia virus-derived reverse transcriptase (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen, Grand Island, NY).

Article Title: A Promising Therapeutic Approach for Multiple Sclerosis: Recombinant T-Cell Receptor Ligands Modulate Experimental Autoimmune Encephalomyelitis by Reducing Interleukin-17 Production and Inhibiting Migration of Encephalitogenic Cells into the CNS
Article Snippet: Alternatively, total RNA was isolated from sorted GFP+ cells using the RNeasy mini-kit protocol (Qiagen) and converted to cDNA using oligo(dT), random hexamers, and Superscript RT II enzyme (Invitrogen, Grand Island, NY). .. Real-time PCR for IL-17 and TNFα was performed using Quantitect SYBR Green PCR master mix (Qiagen) and primers (synthesized by Applied Biosystems, Foster City, CA).

RNA Extraction:

Article Title: Testosterone plays a permissive role in angiotensin II-induced hypertension and cardiac hypertrophy in male rats
Article Snippet: Left ventricular tissues were processed for the total RNA extraction (TRIZOL, Invitrogen, Carlsbad, CA). .. One microgram of total RNA was reverse transcribed using a modified Maloney murine leukemia virus-derived RT (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen).

Article Title: Evolution of the HIV-1 Rev Response Element during Natural Infection Reveals Nucleotide Changes That Correlate with Altered Structure and Increased Activity over Time
Article Snippet: Paragraph title: Viral RNA extraction and cDNA synthesis. ... Briefly, each viral RNA pellet was resuspended in 40 μl of 5 mM Tris-HCl, pH 8.0, prior to cDNA synthesis with 50 μM oligo(dT) and 100 U of SuperScript III reverse transcriptase per reaction mixture (Invitrogen, Carlsbad, CA).

RNA Expression:

Article Title: Influence of hypoxia-inducible factor 1α on dendritic cell differentiation and migration
Article Snippet: First-strand cDNA was synthesized from 1 μg of total RNA using a mixture of oligo(dT) and Revert Aid reverse transcriptase (Fermentas, St.Leon-Rot, Germany). .. RNA expression levels were normalized to β-actin and displayed as fold change relative to normoxic, unstimulated HIF1αflox cells used as calibrator (set to 1).

Agarose Gel Electrophoresis:

Article Title: Involvement of estradiol-17β and its membrane receptor, G protein coupled receptor 30 (GPR30) in regulation of oocyte maturation in zebrafish, Danio rerio
Article Snippet: The integrity of mRNA was confirmed by electrophoresis on a denaturing agarose gel followed by staining with ethidium bromide. .. Total RNA (1–3 μg) was reverse transcribed into cDNA at 50°C for 1 h in 10 μl reaction solution containing 1 × First Strand Buffer, 10 mM dithiothreitol, 0.5 mM each dNTP, 0.5 μg oligo-dT, and 100 U SuperScript III (Invitrogen).

Electrophoresis:

Article Title: Involvement of estradiol-17β and its membrane receptor, G protein coupled receptor 30 (GPR30) in regulation of oocyte maturation in zebrafish, Danio rerio
Article Snippet: The integrity of mRNA was confirmed by electrophoresis on a denaturing agarose gel followed by staining with ethidium bromide. .. Total RNA (1–3 μg) was reverse transcribed into cDNA at 50°C for 1 h in 10 μl reaction solution containing 1 × First Strand Buffer, 10 mM dithiothreitol, 0.5 mM each dNTP, 0.5 μg oligo-dT, and 100 U SuperScript III (Invitrogen).

Quantitation Assay:

Article Title: Inhibiting pyrimidine biosynthesis impairs Ebola virus replication through depletion of nucleoside pools and activation of innate immune responses
Article Snippet: One microgram of total RNA was subjected to cDNA synthesis using oligo(dT) using the SuperScript III first strand cDNA Synthesis Kit following manufacturer’s recommendations (Life Technologies). .. For quantitation of gene expression, real-time PCR with the 2−-ΔΔCT method was used in a total of 20 μl per well with 1:10-fold-diluted cDNA.

Article Title: DRUGS THAT TARGET SPECIFICITY PROTEINS DOWNREGULATE EPIDERMAL GROWTH FACTOR RECEPTOR IN BLADDER CANCER CELLS
Article Snippet: RNA was reverse-transcribed using Superscript II reverse transcriptase (Invitrogen) according to the manufacturer’s protocol. cDNA was prepared from the 253JB-V and KU7 bladder cancer cell lines at different time intervals using a combination of oligodeoxythymidylic acid and dNTP mix (Applied Biosystems, Foster City, CA) and Superscript II. .. The comparative CT method was used for relative quantitation of samples.

Spectrophotometry:

Article Title: Testosterone plays a permissive role in angiotensin II-induced hypertension and cardiac hypertrophy in male rats
Article Snippet: Total RNA concentration and integrity were determined using an ND-1000 Nanodrop spectrophotometer (Thermo Fisher Scientific, Newark, DE). .. One microgram of total RNA was reverse transcribed using a modified Maloney murine leukemia virus-derived RT (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen).

Article Title: Pregnancy upregulates angiotensin type 2 receptor expression and increases blood flow in uterine arteries of rats
Article Snippet: Total RNA concentration and purity were determined using an ND-1000 Nanodrop spectrophotometer (Thermo Fisher Scientific, Newark, DE). .. Two micrograms of total RNA were reverse transcribed using a modified Maloney murine leukemia virus-derived reverse transcriptase (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen, Grand Island, NY).

Concentration Assay:

Article Title: Testosterone plays a permissive role in angiotensin II-induced hypertension and cardiac hypertrophy in male rats
Article Snippet: Total RNA concentration and integrity were determined using an ND-1000 Nanodrop spectrophotometer (Thermo Fisher Scientific, Newark, DE). .. One microgram of total RNA was reverse transcribed using a modified Maloney murine leukemia virus-derived RT (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen).

Article Title: A glycan shield on chimpanzee CD4 protects against infection by primate lentiviruses (HIV/SIV)
Article Snippet: Extracted RNA was used as a template for oligo(dT) primed reverse transcription (SuperScript III RT; Thermo Fisher). .. PCR amplification of CD4 was performed using Phusion High-Fidelity PCR Master Mix (New England Biolabs; #M0531S) containing ∼50 ng of cDNA and 0.2-μM final concentration of each PCR primer CD4-5′ UTR forward 5′-GGGCAAGAAAGACGCAAGC-3′ and CD4-3′ UTR reverse 5′-ATCTGCCTGGCCTCGTG-3′ in a final volume of 50 μL.

Article Title: Pregnancy upregulates angiotensin type 2 receptor expression and increases blood flow in uterine arteries of rats
Article Snippet: Total RNA concentration and purity were determined using an ND-1000 Nanodrop spectrophotometer (Thermo Fisher Scientific, Newark, DE). .. Two micrograms of total RNA were reverse transcribed using a modified Maloney murine leukemia virus-derived reverse transcriptase (New England Biolabs Inc, Ipswich, MA) and a blend of oligo (dT) and random hexamer primers (Invitrogen, Grand Island, NY).

Article Title: Synthesis and Evaluation of Esterified Hsp70 Agonists in Cellular Models of Protein Aggregation and Folding
Article Snippet: RNA concentration and purity were determined by measuring the A260/280 . .. To synthesize cDNA, 0.5 μg of each sample was combined with oligo dT (Thermo Scientific: 25 ng/uL) and incubated at 80oC for 3 minutes.

CTG Assay:

Article Title: Functional Expression of the Heteromeric “Olfactory” Cyclic Nucleotide-Gated Channel in the Hippocampus: A Potential Effector of Synaptic Plasticity in Brain Neurons
Article Snippet: First-strand cDNA was primed with oligo-dT from 25 μg of total RNA pretreated with DNaseI. cDNA was synthesized using the SuperScript II enzyme (Life Technologies) in 50 μl reactions at 42°C for 2 hr. .. The rOCNC2 intron primer sequence is 5′-CAG AAG GCA AGC ACT GAA TGA (GenBank ), and this was used together with a cDNA primer (5′-GGC CAC CAG GTA ACT GTG CTG) to test for the presence of genomic DNA.

Staining:

Article Title: Involvement of estradiol-17β and its membrane receptor, G protein coupled receptor 30 (GPR30) in regulation of oocyte maturation in zebrafish, Danio rerio
Article Snippet: The integrity of mRNA was confirmed by electrophoresis on a denaturing agarose gel followed by staining with ethidium bromide. .. Total RNA (1–3 μg) was reverse transcribed into cDNA at 50°C for 1 h in 10 μl reaction solution containing 1 × First Strand Buffer, 10 mM dithiothreitol, 0.5 mM each dNTP, 0.5 μg oligo-dT, and 100 U SuperScript III (Invitrogen).

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