mmessage mmachine t7 kit  (Thermo Fisher)


Bioz Verified Symbol Thermo Fisher is a verified supplier  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Name:
    mMESSAGE mMACHINE T7 Transcription Kit
    Description:
    mMESSAGE mMACHINE kits are designed for the in vitro synthesis of large amounts of capped RNA. Capped RNA mimics most eukaryotic mRNAs found in vivo, because it has a 7-methyl guanosine cap structure at the 5' end. mMESSAGE mMACHINE kit reactions include cap analog [m7G(5')ppp(5')G] in an ultra high-yield transcription reaction. The cap analog is incorporated only as the first or 5' terminal G of the transcript because its structure precludes its incorporation at any other position in the RNA molecule.How mMESSAGE mMACHINE kits workmMESSAGE mMACHINE kits have a simplified reaction format in which all four ribonucleotides and cap analog are mixed in a single solution. The cap analog:GTP ratio of this solution is 4:1, which is optimal for maximizing both RNA yield and the proportion of capped transcripts. mMESSAGE mMACHINE kits are ideal for the routine synthesis of capped RNAs for oocyte microinjection, in vitro translation, transfection, and other applications. The high yields are achieved by optimizing reaction conditions for RNA synthesis in the presence of high nucleotide concentrations. In addition, the RNA synthesized is protected from degradation by any contaminating ribonucleases that may be present with RNase inhibitor—a component of the Enzyme Mix.Using mMESSAGE mMACHINE KitsIn a 20 µL reaction during a 2 hour incubation, mMESSAGE mMACHINE High Yield Capped RNA Transcription will yield large mass amounts of capped RNA. Up to 10–50 times the yield obtained with conventional in vitro transcription reactions. The ratio of cap analog to GTP has been optimized to allow the best compromise between yield (15–35 µg) and proportion of transcripts that are capped (80%). mMESSAGE mMACHINE Kits also contain a LiCl precipitation solution that is efficient for separating proteins and unincorporated nucleotides (including free cap analog) from the capped RNA, allowing an increased efficiency of translation. mMESSAGE mMACHINE kits are only optimized for use with the polymerases included in the kit. Using a different polymerase may result in low yields. The mMESSAGE mMACHINE kits contain all the buffers and reagents necessary for 25 transcription reactions. Using the control template supplied with the kits (Xenopus elongation factor 1α, pTRI Xef), each mMESSAGE mMACHINE reaction will yield approximately 20–30 µg of RNA using T3 or T7 RNA polymerase, or about 15–25 µg RNA using SP6 RNA polymerase.
    Catalog Number:
    AM1344
    Price:
    None
    Applications:
    In Vitro Transcription|Gene Expression Analysis & Genotyping
    Size:
    25 reactions
    Category:
    Kits and Assays, DNA⁄RNA Detection & Analysis Kits, Transcription Kits
    Score:
    85
    Buy from Supplier
    Name:
    mMESSAGE mMACHINE T7 ULTRA Transcription Kit
    Description:
    The mMESSAGE mMACHINE T7 Ultra Kit combines a new cap analog, anti-reverse cap analog (ARCA), with a patented high-yield transcription technology to generate RNA transcripts that produce higher protein yields compared to other transcripts upon translation. Advantages of the mMESSAGE mMACHINE T7 Ultra Kit:• Synthesize more protein from in vitro-transcribed RNA• Express RNA transcripts more efficiently both in vitro and in vivo• Stabilize transcripts in vivo with included reagents for poly(A) tailingARCA-capped RNA is translationally more activeA base modification in ARCA results in its incorporation in the functional, translatable orientation only; traditional cap analog can be incorporated in both functional and nonfunctional orientations. As a result, incorporating ARCA into transcription reactions yields capped RNAs that are 100% translatable. This is further enhanced by the inclusion of poly(A) tails in mMESSAGE mMACHINE T7 Ultra transcripts. Experiments comparing ARCA and ARCA/poly(A)-tailed transcripts to cap analog and cap analog/poly(A)-tailed transcripts indicate higher levels of protein synthesis with ARCA capped RNA .What Is ARCA?ARCA is a modified cap analog in which the 3' OH group (closer to m7G) is replaced with OCH3 (see schematic). Because of this substitution, the RNA polymerase can only initiate transcription with the remaining hydroxyl group, thus forcing ARCA incorporation in the forward orientation. As a result, 100% of the transcripts synthesized with ARCA at the 5' end are translatable, leading to a strong stimulatory effect on translation.Proper capping of in vitro transcribed RNAProper capping of RNA promotes correct initiation of protein synthesis, as well as stability and processing of mRNA in vivo. Uncapped RNA is rapidly degraded by cellular RNases after microinjection or transfection into cells. Capped RNA is also typically translated more efficiently in in vitro translation systems, generating RNAs with cap analog incorporated only in the functional orientation. Therefore, substitution of traditional cap analog with ARCA results in the synthesis of capped RNAs that are 100% translatable.
    Catalog Number:
    AM1345
    Price:
    None
    Applications:
    In Vitro Transcription|Gene Expression Analysis & Genotyping
    Size:
    10 reactions
    Category:
    Kits and Assays, DNA⁄RNA Detection & Analysis Kits, Transcription Kits
    Score:
    85
    Buy from Supplier


    Structured Review

    Thermo Fisher mmessage mmachine t7 kit
    eGFP ssRNA species produced in vitro . ssRNAs were synthesised from PCR-derived amplicons with a T7 Pol promoter introduced at the 5′ end to facilitate in vitro transcription. PCR amplicons were digested with Bsm BI to define the 3′ end of the transcription cassette. Templates, either 500 ng or 1 µg, were incubated with T7 Pol in the absence or presence of a cap analogue using MEGAscript® or <t>Mmessage</t> <t>Mmachine®.</t> Uncapped ssRNAs were purified and post-transcriptionally capped using ScriptCap™ m7G Capping System. ssRNA was polyadenylated using E. coli polyadenylation polymerase (ePAP) Ambion. Lane R: RiboRuler™ High Range; lanes 1–6: approximately 250 ng each of eGFP ssRNA; uncapped, post-capped, uncapped polyadenylated, post-capped polyadenylated, co-capped, co-capped polyadenylated, respectively. 1.5% TBE AGE 60 V for –90 min. The polyadenylated RNA bands are less sharp as the molecules differ in the numbers of A residues added at the 3′end.
    The mMESSAGE mMACHINE T7 Ultra Kit combines a new cap analog, anti-reverse cap analog (ARCA), with a patented high-yield transcription technology to generate RNA transcripts that produce higher protein yields compared to other transcripts upon translation. Advantages of the mMESSAGE mMACHINE T7 Ultra Kit:• Synthesize more protein from in vitro-transcribed RNA• Express RNA transcripts more efficiently both in vitro and in vivo• Stabilize transcripts in vivo with included reagents for poly(A) tailingARCA-capped RNA is translationally more activeA base modification in ARCA results in its incorporation in the functional, translatable orientation only; traditional cap analog can be incorporated in both functional and nonfunctional orientations. As a result, incorporating ARCA into transcription reactions yields capped RNAs that are 100% translatable. This is further enhanced by the inclusion of poly(A) tails in mMESSAGE mMACHINE T7 Ultra transcripts. Experiments comparing ARCA and ARCA/poly(A)-tailed transcripts to cap analog and cap analog/poly(A)-tailed transcripts indicate higher levels of protein synthesis with ARCA capped RNA .What Is ARCA?ARCA is a modified cap analog in which the 3' OH group (closer to m7G) is replaced with OCH3 (see schematic). Because of this substitution, the RNA polymerase can only initiate transcription with the remaining hydroxyl group, thus forcing ARCA incorporation in the forward orientation. As a result, 100% of the transcripts synthesized with ARCA at the 5' end are translatable, leading to a strong stimulatory effect on translation.Proper capping of in vitro transcribed RNAProper capping of RNA promotes correct initiation of protein synthesis, as well as stability and processing of mRNA in vivo. Uncapped RNA is rapidly degraded by cellular RNases after microinjection or transfection into cells. Capped RNA is also typically translated more efficiently in in vitro translation systems, generating RNAs with cap analog incorporated only in the functional orientation. Therefore, substitution of traditional cap analog with ARCA results in the synthesis of capped RNAs that are 100% translatable.
    https://www.bioz.com/result/mmessage mmachine t7 kit/product/Thermo Fisher
    Average 99 stars, based on 307 article reviews
    Price from $9.99 to $1999.99
    mmessage mmachine t7 kit - by Bioz Stars, 2019-10
    99/100 stars

    Images

    1) Product Images from "Experimental Pathways towards Developing a Rotavirus Reverse Genetics System: Synthetic Full Length Rotavirus ssRNAs Are Neither Infectious nor Translated in Permissive Cells"

    Article Title: Experimental Pathways towards Developing a Rotavirus Reverse Genetics System: Synthetic Full Length Rotavirus ssRNAs Are Neither Infectious nor Translated in Permissive Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0074328

    eGFP ssRNA species produced in vitro . ssRNAs were synthesised from PCR-derived amplicons with a T7 Pol promoter introduced at the 5′ end to facilitate in vitro transcription. PCR amplicons were digested with Bsm BI to define the 3′ end of the transcription cassette. Templates, either 500 ng or 1 µg, were incubated with T7 Pol in the absence or presence of a cap analogue using MEGAscript® or Mmessage Mmachine®. Uncapped ssRNAs were purified and post-transcriptionally capped using ScriptCap™ m7G Capping System. ssRNA was polyadenylated using E. coli polyadenylation polymerase (ePAP) Ambion. Lane R: RiboRuler™ High Range; lanes 1–6: approximately 250 ng each of eGFP ssRNA; uncapped, post-capped, uncapped polyadenylated, post-capped polyadenylated, co-capped, co-capped polyadenylated, respectively. 1.5% TBE AGE 60 V for –90 min. The polyadenylated RNA bands are less sharp as the molecules differ in the numbers of A residues added at the 3′end.
    Figure Legend Snippet: eGFP ssRNA species produced in vitro . ssRNAs were synthesised from PCR-derived amplicons with a T7 Pol promoter introduced at the 5′ end to facilitate in vitro transcription. PCR amplicons were digested with Bsm BI to define the 3′ end of the transcription cassette. Templates, either 500 ng or 1 µg, were incubated with T7 Pol in the absence or presence of a cap analogue using MEGAscript® or Mmessage Mmachine®. Uncapped ssRNAs were purified and post-transcriptionally capped using ScriptCap™ m7G Capping System. ssRNA was polyadenylated using E. coli polyadenylation polymerase (ePAP) Ambion. Lane R: RiboRuler™ High Range; lanes 1–6: approximately 250 ng each of eGFP ssRNA; uncapped, post-capped, uncapped polyadenylated, post-capped polyadenylated, co-capped, co-capped polyadenylated, respectively. 1.5% TBE AGE 60 V for –90 min. The polyadenylated RNA bands are less sharp as the molecules differ in the numbers of A residues added at the 3′end.

    Techniques Used: Produced, In Vitro, Polymerase Chain Reaction, Derivative Assay, Incubation, Purification

    Related Articles

    Clone Assay:

    Article Title: An allosteric link connecting the lipid-protein interface to the gating of the nicotinic acetylcholine receptor
    Article Snippet: Wild-type human α1 , β1 , δ, and ε nAChR-pRBG4 clones were kindly provided by Steven Sine. .. The α1 , β1 , δ, and ε nAChR-pcDNA3 was linearized with XhoI and capped cRNA was produced by in vitro transcription using the mMESSAGE mMACHINE® T7 kit (Ambion).

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: The control firefly luciferase gene was amplified from the plasmid pGL3-basic (Promega GmbH, Mannheim, Germany) and cloned into pT7CFE1-cMyc at XhoI and NotI restriction sites. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol.

    Article Title: A missense mutation in ASRGL1 is involved in causing autosomal recessive retinal degeneration
    Article Snippet: The efficacy of MO injections was evaluated by semi-quantitative RT-PCR using RNA extracted from injected embryos at 1 dpf and using primer sets to amplify Asrgl1 transcript Asrgl1startsite_MO Fwd, 5′- GAAAGAGGCAGCCAGGACTG-3′ and Asrgl1startsite_MO Rev, 5′- CAATGCATCCATCTCTACCTC-3′; Asrgl1 splice-site_MO Fwd, 5′- GGAAGTGCCCGAGGAGTCATT-3′ and Asrgl1 splice-site_MO Rev 5′- CTTCTCCGTGGCCTGTGGG-3′. .. The recombinant wt-h ASRGL1 and G178R -h ASRGL1 clones were used to generate the full-length mRNA with the mMESSAGE mMACHINE T7 kit (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions. .. The mRNA was analyzed for size and quality using a Bioanalyzer RNA nano Chip (Agilent, Santa Clara, CA, USA).

    Luciferase:

    Article Title: miR-122 does not impact recognition of the HCV genome by innate sensors of RNA but rather protects the 5′ end from the cellular pyrophosphatases, DOM3Z and DUSP11
    Article Snippet: Firefly luciferase mRNA was transcribed from the Luciferase T7 Control DNA plasmid (Promega), linearized using XmnI , while Renilla luciferase mRNA was transcribed from the pRL-TK plasmid (Promega), linearized using BglII . .. Both luciferase messenger RNAs were in vitro transcribed using the mMessage mMachine T7 Kit (Life Technologies) according to the manufacturer's protocol. .. The triple-FLAG-tagged (3xF) IFIT1, IFIT5 and the empty vector ( ) were kindly provided by Kathleen Collins (UC Berkeley).

    Article Title: Spicatoside A derived from Liriope platyphylla root ethanol extract inhibits hepatitis E virus genotype 3 replication in vitro
    Article Snippet: Paragraph title: HEV replicon, in vitro transcription, transfection and luciferase-reporter assay ... Capped RNA transcripts from pSHEV3-luc and luc-pcDNA3 plasmids were generated using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific, Waltham, MA) according to the manufacturer’s instructions.

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: The control firefly luciferase gene was amplified from the plasmid pGL3-basic (Promega GmbH, Mannheim, Germany) and cloned into pT7CFE1-cMyc at XhoI and NotI restriction sites. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol.

    Reporter Assay:

    Article Title: Spicatoside A derived from Liriope platyphylla root ethanol extract inhibits hepatitis E virus genotype 3 replication in vitro
    Article Snippet: Capped RNA transcripts from pSHEV3-luc and luc-pcDNA3 plasmids were generated using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific, Waltham, MA) according to the manufacturer’s instructions. .. Huh 7.5 cells were transfected with capped RNA transcripts produced from pSHEV3-luc (0.34 μg/104 cells) and luc-pcDNA3 (0.17 μg/104 cells) plasmids using DMRIE-C reagent (Invitrogen, Carlsbad, CA) in keeping with the manufacturer’s instructions.

    Synthesized:

    Article Title: Targeting fidelity of adenine and cytosine base editors in mouse embryos
    Article Snippet: The sgRNAs were designed using CRISPR.MIT.EDU, and subsequently produced using ThermoFisher’s sgRNA In Vitro Transcription Service. .. ABE, BE4, and VQR-BE3 mRNAs were synthesized in vitro using the mMESSAGE mMACHINE T7 kit (ThermoFisher Scientific). .. Deaminase fused-Cas9 mRNA (50 ng/μl for each base editor) and sgRNAs (20 ng/μl for each sgRNA) were mixed and co-microinjected into the cytoplasm of fertilized eggs collected from superovulated C57BL/6N female mice (Charles River Laboratories) and implanted into oviducts of pseudopregnant fosters (Swiss Webster).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. Purified cRNAs were resuspended in nuclease-free water at a concentration of 0.2 μg/μl and stored at −80°C.

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. Purified cRNAs were resuspended in nuclease-free water at a concentration of 0.2 μg/μl and stored at −80°C.

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: Tmem63c cDNA (Rattus norvegicus) was synthesized by Thermo Fisher Scientific using their GeneArt Gene synthesis service. .. In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Far Upstream Element-Binding Protein 1 Binds the 3′ Untranslated Region of PKD2 and Suppresses Its Translation
    Article Snippet: The MO sequences were as follows: pkd2 , 5′-AGGACGAACGCGACTGGAGCTCATC-3′; fubp1 , 5′- GGCCATGTCTGCACGAACAGTCTTC-3′; gli2 (an antisense mismatch morpholino, used as a negative control), 5′-CCTCTTACCTCAGTTACAATTTATA-3′. .. Capped human fubp1 mRNA was synthesized using the mMessage mMachine T7 kit (Ambion, Austin, TX) and injected into fertilized embryos (at 200 pg each). .. Embryos at 3 dpf were fixed in 1.5% glutaraldehyde, 1% paraformaldehyde, 70 mM sodium phosphate, pH 7.2, and 3% sucrose overnight at 4°C.

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: Alanine-coding mutations for conserved and control residues were introduced into full-length cDNA clone pEAV211 ( ) using appropriate shuttle vectors and restriction enzymes. .. The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Virus replication was monitored by immunofluorescence microscopy until 72 h post transfection (p.t.) using antibodies directed against nsp3 and N protein as described ( ) and by plaque assays ( ) using transfected cell culture supernatants, to monitor the production of viral progeny.

    Quantitative RT-PCR:

    Article Title: Recombinant Porcine Reproductive and Respiratory Syndrome Virus Expressing Membrane-Bound Interleukin-15 as an Immunomodulatory Adjuvant Enhances NK and γδ T Cell Responses and Confers Heterologous Protection
    Article Snippet: The quantification of PRRSV RNA copy numbers in sera and in lung tissues was performed by qRT-PCR using a SYBR green one-step qRT-PCR kit (Bioline), as described previously ( , ). .. The RNA standard used for qRT-PCR was derived from the in vitro transcription of a PRRSV full-length cDNA clone, pACYC-VR2385, by using the mMessage mMachine T7 kit (Ambion). .. Each qRT-PCR was performed in triplicate.

    SYBR Green Assay:

    Article Title: Recombinant Porcine Reproductive and Respiratory Syndrome Virus Expressing Membrane-Bound Interleukin-15 as an Immunomodulatory Adjuvant Enhances NK and γδ T Cell Responses and Confers Heterologous Protection
    Article Snippet: The quantification of PRRSV RNA copy numbers in sera and in lung tissues was performed by qRT-PCR using a SYBR green one-step qRT-PCR kit (Bioline), as described previously ( , ). .. The RNA standard used for qRT-PCR was derived from the in vitro transcription of a PRRSV full-length cDNA clone, pACYC-VR2385, by using the mMessage mMachine T7 kit (Ambion).

    Incubation:

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. After being cultured overnight at 18°C, oocytes were microinjected with 27.6 nl Cs Drip1 cRNA (5.52 ng) and 27.6 nl nuclease-free water as control.

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. After being cultured overnight at 18°C, oocytes were microinjected with 27.6 nl Cs Drip1 cRNA (5.52 ng) and 27.6 nl nuclease-free water as control.

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol. .. In brief, the original pT7CFE1-cMyc plasmid (for non-coding (NC) mRNA control) or the plasmid carrying spa , mecA , sitC or luc gene was linearized.

    Amplification:

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: Firstly, the entire coding region of Cs Drip1 was amplified with a high-fidelity polymerase (PrimeSTAR®, HS DNA polymerase. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: Firstly, the entire coding region of Cs Drip1 was amplified with a high-fidelity polymerase (PrimeSTAR®, HS DNA polymerase. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: The control firefly luciferase gene was amplified from the plasmid pGL3-basic (Promega GmbH, Mannheim, Germany) and cloned into pT7CFE1-cMyc at XhoI and NotI restriction sites. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol.

    Article Title: Unraveling the transcriptional regulation of TWIST1 in limb development
    Article Snippet: The PCR amplified T7-sgRNA product was used as template for in vitro transcription using the MEGAshortscript T7 kit (Thermo Fisher Scientific). .. Cas9 mRNA was transcribed in vitro using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific).

    Activity Assay:

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol.

    Infection:

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Sequence analysis of the nsp9-coding region was performed to either verify the presence of the introduced mutations or to monitor the presence of (second site) reversions.

    Expressing:

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction. .. The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction.

    Article Title: An allosteric link connecting the lipid-protein interface to the gating of the nicotinic acetylcholine receptor
    Article Snippet: Paragraph title: cRNA constructs for oocyte expression ... The α1 , β1 , δ, and ε nAChR-pcDNA3 was linearized with XhoI and capped cRNA was produced by in vitro transcription using the mMESSAGE mMACHINE® T7 kit (Ambion).

    Modification:

    Article Title: Unraveling the transcriptional regulation of TWIST1 in limb development
    Article Snippet: Generation of eTw5-7 deletion mice using CRISPR/Cas9 A mouse strain carrying deleted eTw5-7 alleles were created using a modified CRISPR/Cas9 protocol [ ]. .. Cas9 mRNA was transcribed in vitro using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific).

    Over Expression:

    Article Title: A missense mutation in ASRGL1 is involved in causing autosomal recessive retinal degeneration
    Article Snippet: Paragraph title: Overexpression of hASRGL1 in zebrafish ... The recombinant wt-h ASRGL1 and G178R -h ASRGL1 clones were used to generate the full-length mRNA with the mMESSAGE mMACHINE T7 kit (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions.

    Derivative Assay:

    Article Title: Recombinant Porcine Reproductive and Respiratory Syndrome Virus Expressing Membrane-Bound Interleukin-15 as an Immunomodulatory Adjuvant Enhances NK and γδ T Cell Responses and Confers Heterologous Protection
    Article Snippet: The quantification of PRRSV RNA copy numbers in sera and in lung tissues was performed by qRT-PCR using a SYBR green one-step qRT-PCR kit (Bioline), as described previously ( , ). .. The RNA standard used for qRT-PCR was derived from the in vitro transcription of a PRRSV full-length cDNA clone, pACYC-VR2385, by using the mMessage mMachine T7 kit (Ambion). .. Each qRT-PCR was performed in triplicate.

    Article Title: miR-122 does not impact recognition of the HCV genome by innate sensors of RNA but rather protects the 5′ end from the cellular pyrophosphatases, DOM3Z and DUSP11
    Article Snippet: Plasmids pJ6/JFH-1 FL Rluc WT and pJ6/JFH-1 FL Rluc GNN bear full-length viral sequences derived from the J6 (structural proteins) and JFH-1 (non-structural proteins) isolates of HCV, and a Renilla luciferase reporter ( ). .. Both luciferase messenger RNAs were in vitro transcribed using the mMessage mMachine T7 Kit (Life Technologies) according to the manufacturer's protocol.

    Electroporation:

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: DENV2 16681 strain RNA was transcribed in vitro using mMessage/mMachine T7 kit (Ambion) from pD2IC-30P-NBX plasmid linearized by XbaI. .. A Renilla reporter DENV2 NGC strain RNA was transcribed in vitro by mMessage/mMachine T7 kit (Ambion) from pACYC-Rluc2A-NGC linearized by XbaI.

    Transfection:

    Article Title: miR-122 does not impact recognition of the HCV genome by innate sensors of RNA but rather protects the 5′ end from the cellular pyrophosphatases, DOM3Z and DUSP11
    Article Snippet: Both luciferase messenger RNAs were in vitro transcribed using the mMessage mMachine T7 Kit (Life Technologies) according to the manufacturer's protocol. .. The triple-FLAG-tagged (3xF) IFIT1, IFIT5 and the empty vector ( ) were kindly provided by Kathleen Collins (UC Berkeley).

    Article Title: Spicatoside A derived from Liriope platyphylla root ethanol extract inhibits hepatitis E virus genotype 3 replication in vitro
    Article Snippet: Paragraph title: HEV replicon, in vitro transcription, transfection and luciferase-reporter assay ... Capped RNA transcripts from pSHEV3-luc and luc-pcDNA3 plasmids were generated using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific, Waltham, MA) according to the manufacturer’s instructions.

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol.

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: DENV was produced by transfection of viral RNA into Huh7 cells and harvesting the culture supernatants at days 5–7. .. A Renilla reporter DENV2 NGC strain RNA was transcribed in vitro by mMessage/mMachine T7 kit (Ambion) from pACYC-Rluc2A-NGC linearized by XbaI.

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: Alanine-coding mutations for conserved and control residues were introduced into full-length cDNA clone pEAV211 ( ) using appropriate shuttle vectors and restriction enzymes. .. The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Virus replication was monitored by immunofluorescence microscopy until 72 h post transfection (p.t.) using antibodies directed against nsp3 and N protein as described ( ) and by plaque assays ( ) using transfected cell culture supernatants, to monitor the production of viral progeny.

    Cell Culture:

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction. .. The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction.

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Sequence analysis of the nsp9-coding region was performed to either verify the presence of the introduced mutations or to monitor the presence of (second site) reversions.

    Generated:

    Article Title: Spicatoside A derived from Liriope platyphylla root ethanol extract inhibits hepatitis E virus genotype 3 replication in vitro
    Article Snippet: The transfection control firefly luciferase-pcDNA3 (luc-pcDNA3) plasmid was a gift from William Kaelin (Addgene plasmid # 18964) . pSHEV3-luc plasmids were linearized by treatment with Xba I (New England BioLabs, Ipswich, MA) and firefly luciferase plasmid linearized with Bgl II (New England BioLabs). .. Capped RNA transcripts from pSHEV3-luc and luc-pcDNA3 plasmids were generated using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific, Waltham, MA) according to the manufacturer’s instructions. .. Huh 7.5 cells were transfected with capped RNA transcripts produced from pSHEV3-luc (0.34 μg/104 cells) and luc-pcDNA3 (0.17 μg/104 cells) plasmids using DMRIE-C reagent (Invitrogen, Carlsbad, CA) in keeping with the manufacturer’s instructions.

    Article Title: A missense mutation in ASRGL1 is involved in causing autosomal recessive retinal degeneration
    Article Snippet: The recombinant wt-h ASRGL1 and G178R -h ASRGL1 clones were used to generate the full-length mRNA with the mMESSAGE mMACHINE T7 kit (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions. .. Each embryo was injected with a specific dose of mRNA (ranging from 1 to 200 pg) of wt-h ASRGL1 or G178R-h ASRGL1 mRNA and/or phenol red dye.

    Gel Extraction:

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments tmem63c cDNA (Danio rerio ) was linearized using ApaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments, Tmem63c cDNA was linearized using XbaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Polymerase Chain Reaction:

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Unraveling the transcriptional regulation of TWIST1 in limb development
    Article Snippet: The PCR amplified T7-sgRNA product was used as template for in vitro transcription using the MEGAshortscript T7 kit (Thermo Fisher Scientific). .. Cas9 mRNA was transcribed in vitro using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific).

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. For this purpose, fresh BHK-21 cells were infected with virus-containing cell culture supernatants and total RNA was extracted with Tripure Isolation Reagent (Roche Applied Science) after appearance of cytopathic effect (CPE) (typically at 18 h post infection (p.i.)).

    Injection:

    Article Title: Hydrophobic pore gates regulate ion permeation in polycystic kidney disease 2 and 2L1 channels
    Article Snippet: PKD2 was knocked down in zebrafish by injection of a translation-blocking antisense morpholino oligonucleotide (MO) (Gene Tools LLC, Philomath, OR) into fertilized eggs within 1 h postfertilization (at 2.5 ng each). .. Capped mRNAs of human PKD2 WT or mutants were in vitro transcripted with mMessage mMachine T7 kit (Ambion) and co-injected into fertilized eggs with PKD2 MO (at 100 pg each).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. After being cultured overnight at 18°C, oocytes were microinjected with 27.6 nl Cs Drip1 cRNA (5.52 ng) and 27.6 nl nuclease-free water as control.

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. After being cultured overnight at 18°C, oocytes were microinjected with 27.6 nl Cs Drip1 cRNA (5.52 ng) and 27.6 nl nuclease-free water as control.

    Article Title: A missense mutation in ASRGL1 is involved in causing autosomal recessive retinal degeneration
    Article Snippet: The recombinant wt-h ASRGL1 and G178R -h ASRGL1 clones were used to generate the full-length mRNA with the mMESSAGE mMACHINE T7 kit (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions. .. The recombinant wt-h ASRGL1 and G178R -h ASRGL1 clones were used to generate the full-length mRNA with the mMESSAGE mMACHINE T7 kit (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions.

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion). .. In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion). .. In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Far Upstream Element-Binding Protein 1 Binds the 3′ Untranslated Region of PKD2 and Suppresses Its Translation
    Article Snippet: The MO sequences were as follows: pkd2 , 5′-AGGACGAACGCGACTGGAGCTCATC-3′; fubp1 , 5′- GGCCATGTCTGCACGAACAGTCTTC-3′; gli2 (an antisense mismatch morpholino, used as a negative control), 5′-CCTCTTACCTCAGTTACAATTTATA-3′. .. Capped human fubp1 mRNA was synthesized using the mMessage mMachine T7 kit (Ambion, Austin, TX) and injected into fertilized embryos (at 200 pg each). .. Embryos at 3 dpf were fixed in 1.5% glutaraldehyde, 1% paraformaldehyde, 70 mM sodium phosphate, pH 7.2, and 3% sucrose overnight at 4°C.

    Binding Assay:

    Article Title: miR-122 does not impact recognition of the HCV genome by innate sensors of RNA but rather protects the 5′ end from the cellular pyrophosphatases, DOM3Z and DUSP11
    Article Snippet: Plasmids pSGR p3 S1S2 Fluc WT and pSGR p3 S1S2 Fluc GND bear sub-genomic JFH-1-derived replicons with a firefly luciferase reporter ( ) and have C to G mutations at position 3 in the miR-122 seed binding sites S1 and S2 in the HCV 5′UTR ( ). .. Both luciferase messenger RNAs were in vitro transcribed using the mMessage mMachine T7 Kit (Life Technologies) according to the manufacturer's protocol.

    Immunofluorescence:

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction. .. The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction.

    In Vivo:

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments tmem63c cDNA (Danio rerio ) was linearized using ApaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments, Tmem63c cDNA was linearized using XbaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Isolation:

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Sequence analysis of the nsp9-coding region was performed to either verify the presence of the introduced mutations or to monitor the presence of (second site) reversions.

    Functional Assay:

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: Paragraph title: Functional oocyte swelling assays ... The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol.

    Microscopy:

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction. .. The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction.

    Article Title: Far Upstream Element-Binding Protein 1 Binds the 3′ Untranslated Region of PKD2 and Suppresses Its Translation
    Article Snippet: Capped human fubp1 mRNA was synthesized using the mMessage mMachine T7 kit (Ambion, Austin, TX) and injected into fertilized embryos (at 200 pg each). .. Capped human fubp1 mRNA was synthesized using the mMessage mMachine T7 kit (Ambion, Austin, TX) and injected into fertilized embryos (at 200 pg each).

    Purification:

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments tmem63c cDNA (Danio rerio ) was linearized using ApaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments, Tmem63c cDNA was linearized using XbaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Sequencing:

    Article Title: Hydrophobic pore gates regulate ion permeation in polycystic kidney disease 2 and 2L1 channels
    Article Snippet: The PKD2 MO sequence was: 5′-AGGACGAACGCGACTGGAGCTCATC-3′. .. Capped mRNAs of human PKD2 WT or mutants were in vitro transcripted with mMessage mMachine T7 kit (Ambion) and co-injected into fertilized eggs with PKD2 MO (at 100 pg each).

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: Mutations in the SARS-CoV nsp12-coding region were engineered in prSCV, a pBeloBac11 derivative containing a full-length cDNA copy of the SARS-CoV Frankfurt-1 sequence ( ) by using ‘en passant recombineering’ as described in Tischer et al . ( ). .. The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction.

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: TaKaRa, Tokyo, Japan) using primers with Kozak sequence and restriction enzyme cutting sites (SpeI and NotI). .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: TaKaRa, Tokyo, Japan) using primers with Kozak sequence and restriction enzyme cutting sites (SpeI and NotI). .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Unraveling the transcriptional regulation of TWIST1 in limb development
    Article Snippet: No potential off-targets were found when searching for matches in the mouse genome (mm9) when allowing for up to two mismatches in the 20 nucleotide-long sequence preceding the PAM sequence. .. Cas9 mRNA was transcribed in vitro using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific).

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: Alanine-coding mutations for conserved and control residues were introduced into full-length cDNA clone pEAV211 ( ) using appropriate shuttle vectors and restriction enzymes. .. The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Virus replication was monitored by immunofluorescence microscopy until 72 h post transfection (p.t.) using antibodies directed against nsp3 and N protein as described ( ) and by plaque assays ( ) using transfected cell culture supernatants, to monitor the production of viral progeny.

    Construct:

    Article Title: An allosteric link connecting the lipid-protein interface to the gating of the nicotinic acetylcholine receptor
    Article Snippet: Paragraph title: cRNA constructs for oocyte expression ... The α1 , β1 , δ, and ε nAChR-pcDNA3 was linearized with XhoI and capped cRNA was produced by in vitro transcription using the mMESSAGE mMACHINE® T7 kit (Ambion).

    Cotransfection:

    Article Title: miR-122 does not impact recognition of the HCV genome by innate sensors of RNA but rather protects the 5′ end from the cellular pyrophosphatases, DOM3Z and DUSP11
    Article Snippet: Both luciferase messenger RNAs were in vitro transcribed using the mMessage mMachine T7 Kit (Life Technologies) according to the manufacturer's protocol. .. Plasmid DNA along with viral RNA and miRNAs were transfected using Lipofectamine 2000 according to manufacturer's protocol.

    CRISPR:

    Article Title: Targeting fidelity of adenine and cytosine base editors in mouse embryos
    Article Snippet: The sgRNAs were designed using CRISPR.MIT.EDU, and subsequently produced using ThermoFisher’s sgRNA In Vitro Transcription Service. .. ABE, BE4, and VQR-BE3 mRNAs were synthesized in vitro using the mMESSAGE mMACHINE T7 kit (ThermoFisher Scientific).

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: Paragraph title: Rescue of CRISPR/Cas9-mediated tmem63c somatic mutants and tmem63c ex2-sdMO-mediated gene knockdown ... In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Unraveling the transcriptional regulation of TWIST1 in limb development
    Article Snippet: Paragraph title: Generation of eTw5-7 deletion mice using CRISPR/Cas9 ... Cas9 mRNA was transcribed in vitro using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific).

    Mouse Assay:

    Article Title: Unraveling the transcriptional regulation of TWIST1 in limb development
    Article Snippet: Paragraph title: Generation of eTw5-7 deletion mice using CRISPR/Cas9 ... Cas9 mRNA was transcribed in vitro using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific).

    Plasmid Preparation:

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA).

    Article Title: LY6E mediates an evolutionarily conserved enhancement of virus infection by targeting a late entry step
    Article Snippet: The following viral/replicon RNA were in vitro transcribed with mMessage mMachine SP6 kit (Ambion): YFV-17D-Venus RNA from XhoI-linearized YF17D(5’C25Venus2AUbi) plasmid , YFV-17D from XhoI-linearized pACNR-17D-Yfx , YFV-R.luc2A-RP replicon RNA from XhoI-linearized YFV-R.luc2A-RP . .. The following RNAs were in vitro transcribed with mMessage mMachine T7 kit (Ambion): DENV2-GFP RNA from XbaI-linearized pDV2.IC30P.A.eGFP.P2AUbFIX plasmid . .. RNA was purified from the transcription reaction using RNeasy mini kit (Qiagen) and quantified by Nanodrop.

    Article Title: miR-122 does not impact recognition of the HCV genome by innate sensors of RNA but rather protects the 5′ end from the cellular pyrophosphatases, DOM3Z and DUSP11
    Article Snippet: Firefly luciferase mRNA was transcribed from the Luciferase T7 Control DNA plasmid (Promega), linearized using XmnI , while Renilla luciferase mRNA was transcribed from the pRL-TK plasmid (Promega), linearized using BglII . .. Both luciferase messenger RNAs were in vitro transcribed using the mMessage mMachine T7 Kit (Life Technologies) according to the manufacturer's protocol.

    Article Title: An allosteric link connecting the lipid-protein interface to the gating of the nicotinic acetylcholine receptor
    Article Snippet: Each nAChR subunit DNA was transferred into the pcDNA3 vector as an EcoRI fragment. .. The α1 , β1 , δ, and ε nAChR-pcDNA3 was linearized with XhoI and capped cRNA was produced by in vitro transcription using the mMESSAGE mMACHINE® T7 kit (Ambion).

    Article Title: Spicatoside A derived from Liriope platyphylla root ethanol extract inhibits hepatitis E virus genotype 3 replication in vitro
    Article Snippet: The transfection control firefly luciferase-pcDNA3 (luc-pcDNA3) plasmid was a gift from William Kaelin (Addgene plasmid # 18964) . pSHEV3-luc plasmids were linearized by treatment with Xba I (New England BioLabs, Ipswich, MA) and firefly luciferase plasmid linearized with Bgl II (New England BioLabs). .. Capped RNA transcripts from pSHEV3-luc and luc-pcDNA3 plasmids were generated using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific, Waltham, MA) according to the manufacturer’s instructions.

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: The control firefly luciferase gene was amplified from the plasmid pGL3-basic (Promega GmbH, Mannheim, Germany) and cloned into pT7CFE1-cMyc at XhoI and NotI restriction sites. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol.

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: DENV2 16681 strain RNA was transcribed in vitro using mMessage/mMachine T7 kit (Ambion) from pD2IC-30P-NBX plasmid linearized by XbaI. .. A Renilla reporter DENV2 NGC strain RNA was transcribed in vitro by mMessage/mMachine T7 kit (Ambion) from pACYC-Rluc2A-NGC linearized by XbaI.

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: Alanine-coding mutations for conserved and control residues were introduced into full-length cDNA clone pEAV211 ( ) using appropriate shuttle vectors and restriction enzymes. .. The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Virus replication was monitored by immunofluorescence microscopy until 72 h post transfection (p.t.) using antibodies directed against nsp3 and N protein as described ( ) and by plaque assays ( ) using transfected cell culture supernatants, to monitor the production of viral progeny.

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: Open reading frames (ORFs) encoding 26 hits were selected from the Human ORFeome library of cDNA clones( ) (Open Biosystems), three from Addgene and one from DNASU( ) and recombined into a pFLAG (for FLAG tagging) vector using Gateway technology (Invitrogen). .. DENV2 16681 strain RNA was transcribed in vitro using mMessage/mMachine T7 kit (Ambion) from pD2IC-30P-NBX plasmid linearized by XbaI. .. DENV was produced by transfection of viral RNA into Huh7 cells and harvesting the culture supernatants at days 5–7.

    Negative Control:

    Article Title: Far Upstream Element-Binding Protein 1 Binds the 3′ Untranslated Region of PKD2 and Suppresses Its Translation
    Article Snippet: The MO sequences were as follows: pkd2 , 5′-AGGACGAACGCGACTGGAGCTCATC-3′; fubp1 , 5′- GGCCATGTCTGCACGAACAGTCTTC-3′; gli2 (an antisense mismatch morpholino, used as a negative control), 5′-CCTCTTACCTCAGTTACAATTTATA-3′. .. Capped human fubp1 mRNA was synthesized using the mMessage mMachine T7 kit (Ambion, Austin, TX) and injected into fertilized embryos (at 200 pg each).

    Recombinant:

    Article Title: A missense mutation in ASRGL1 is involved in causing autosomal recessive retinal degeneration
    Article Snippet: The efficacy of MO injections was evaluated by semi-quantitative RT-PCR using RNA extracted from injected embryos at 1 dpf and using primer sets to amplify Asrgl1 transcript Asrgl1startsite_MO Fwd, 5′- GAAAGAGGCAGCCAGGACTG-3′ and Asrgl1startsite_MO Rev, 5′- CAATGCATCCATCTCTACCTC-3′; Asrgl1 splice-site_MO Fwd, 5′- GGAAGTGCCCGAGGAGTCATT-3′ and Asrgl1 splice-site_MO Rev 5′- CTTCTCCGTGGCCTGTGGG-3′. .. The recombinant wt-h ASRGL1 and G178R -h ASRGL1 clones were used to generate the full-length mRNA with the mMESSAGE mMACHINE T7 kit (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions. .. The mRNA was analyzed for size and quality using a Bioanalyzer RNA nano Chip (Agilent, Santa Clara, CA, USA).

    In Vitro:

    Article Title: Experimental Pathways towards Developing a Rotavirus Reverse Genetics System: Synthetic Full Length Rotavirus ssRNAs Are Neither Infectious nor Translated in Permissive Cells
    Article Snippet: Paragraph title: In vitro Transcription of viral Positive sense RNAs from Digested pUC19T7 RV Templates ... For co-transcriptional capping of ssRNAs, the Mmessage Mmachine T7 kit (Ambion) was used with the following alterations: 1.5 µl of T7 RNA Pol (Ambion; 20 u/ul) per reaction was added, and the amount of linear template was increased to 1 µg for each reaction.

    Article Title: Hydrophobic pore gates regulate ion permeation in polycystic kidney disease 2 and 2L1 channels
    Article Snippet: The PKD2 MO sequence was: 5′-AGGACGAACGCGACTGGAGCTCATC-3′. .. Capped mRNAs of human PKD2 WT or mutants were in vitro transcripted with mMessage mMachine T7 kit (Ambion) and co-injected into fertilized eggs with PKD2 MO (at 100 pg each). .. This study has been approved by the Ethical Committee for Animal Experiments (University of Alberta) and was performed according to the Guidelines for Research with Experimental Animals (University of Alberta) and for the Care and Use of Laboratory Animals (NIH guide, revised in 1996).

    Article Title: Targeting fidelity of adenine and cytosine base editors in mouse embryos
    Article Snippet: The sgRNAs were designed using CRISPR.MIT.EDU, and subsequently produced using ThermoFisher’s sgRNA In Vitro Transcription Service. .. ABE, BE4, and VQR-BE3 mRNAs were synthesized in vitro using the mMESSAGE mMACHINE T7 kit (ThermoFisher Scientific). .. Deaminase fused-Cas9 mRNA (50 ng/μl for each base editor) and sgRNAs (20 ng/μl for each sgRNA) were mixed and co-microinjected into the cytoplasm of fertilized eggs collected from superovulated C57BL/6N female mice (Charles River Laboratories) and implanted into oviducts of pseudopregnant fosters (Swiss Webster).

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. Purified cRNAs were resuspended in nuclease-free water at a concentration of 0.2 μg/μl and stored at −80°C.

    Article Title: Identification and Functional Analysis of the First Aquaporin from Striped Stem Borer, Chilo suppressalis
    Article Snippet: And the PCR products were digested with SpeI and NotI, and subcloned into pT7Ts vector (Invitrogen, Carlsbad, California, USA), and then plasmids were fully linearized with SmaI. .. The cRNAs of Cs Drip1 were synthesized in vitro using mMESSAGE mMACHINE T7 kit (Ambion, Austin, TX, USA). .. Purified cRNAs were resuspended in nuclease-free water at a concentration of 0.2 μg/μl and stored at −80°C.

    Article Title: LY6E mediates an evolutionarily conserved enhancement of virus infection by targeting a late entry step
    Article Snippet: The following viral/replicon RNA were in vitro transcribed with mMessage mMachine SP6 kit (Ambion): YFV-17D-Venus RNA from XhoI-linearized YF17D(5’C25Venus2AUbi) plasmid , YFV-17D from XhoI-linearized pACNR-17D-Yfx , YFV-R.luc2A-RP replicon RNA from XhoI-linearized YFV-R.luc2A-RP . .. The following RNAs were in vitro transcribed with mMessage mMachine T7 kit (Ambion): DENV2-GFP RNA from XbaI-linearized pDV2.IC30P.A.eGFP.P2AUbFIX plasmid . .. RNA was purified from the transcription reaction using RNeasy mini kit (Qiagen) and quantified by Nanodrop.

    Article Title: Recombinant Porcine Reproductive and Respiratory Syndrome Virus Expressing Membrane-Bound Interleukin-15 as an Immunomodulatory Adjuvant Enhances NK and γδ T Cell Responses and Confers Heterologous Protection
    Article Snippet: The quantification of PRRSV RNA copy numbers in sera and in lung tissues was performed by qRT-PCR using a SYBR green one-step qRT-PCR kit (Bioline), as described previously ( , ). .. The RNA standard used for qRT-PCR was derived from the in vitro transcription of a PRRSV full-length cDNA clone, pACYC-VR2385, by using the mMessage mMachine T7 kit (Ambion). .. Each qRT-PCR was performed in triplicate.

    Article Title: miR-122 does not impact recognition of the HCV genome by innate sensors of RNA but rather protects the 5′ end from the cellular pyrophosphatases, DOM3Z and DUSP11
    Article Snippet: Firefly luciferase mRNA was transcribed from the Luciferase T7 Control DNA plasmid (Promega), linearized using XmnI , while Renilla luciferase mRNA was transcribed from the pRL-TK plasmid (Promega), linearized using BglII . .. Both luciferase messenger RNAs were in vitro transcribed using the mMessage mMachine T7 Kit (Life Technologies) according to the manufacturer's protocol. .. The triple-FLAG-tagged (3xF) IFIT1, IFIT5 and the empty vector ( ) were kindly provided by Kathleen Collins (UC Berkeley).

    Article Title: An allosteric link connecting the lipid-protein interface to the gating of the nicotinic acetylcholine receptor
    Article Snippet: Each nAChR subunit DNA was transferred into the pcDNA3 vector as an EcoRI fragment. .. The α1 , β1 , δ, and ε nAChR-pcDNA3 was linearized with XhoI and capped cRNA was produced by in vitro transcription using the mMESSAGE mMACHINE® T7 kit (Ambion). .. All mutants were created using QuikChange™ Site-Directed Mutagenesis kits (Agilent) and verified by sequencing.

    Article Title: Spicatoside A derived from Liriope platyphylla root ethanol extract inhibits hepatitis E virus genotype 3 replication in vitro
    Article Snippet: Paragraph title: HEV replicon, in vitro transcription, transfection and luciferase-reporter assay ... Capped RNA transcripts from pSHEV3-luc and luc-pcDNA3 plasmids were generated using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific, Waltham, MA) according to the manufacturer’s instructions.

    Article Title: Antigen delivery to dendritic cells shapes human CD4+ and CD8+ T cell memory responses to Staphylococcus aureus
    Article Snippet: The control firefly luciferase gene was amplified from the plasmid pGL3-basic (Promega GmbH, Mannheim, Germany) and cloned into pT7CFE1-cMyc at XhoI and NotI restriction sites. .. In vitro transcription (ivT) was performed with mMESSAGE mMachine T7 Kit (Ambion,Thermo Fisher, Paisley, UK) following the manufacturer’s protocol. .. In brief, the original pT7CFE1-cMyc plasmid (for non-coding (NC) mRNA control) or the plasmid carrying spa , mecA , sitC or luc gene was linearized.

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments tmem63c cDNA (Danio rerio ) was linearized using ApaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion). .. For poly-A-tailing, the Poly(A)-tailing Kit (Ambion) was used, followed by RNA extraction using RNeasy Mini Kit (Qiagen).

    Article Title: Unraveling the transcriptional regulation of TWIST1 in limb development
    Article Snippet: The PCR amplified T7-sgRNA product was used as template for in vitro transcription using the MEGAshortscript T7 kit (Thermo Fisher Scientific). .. Cas9 mRNA was transcribed in vitro using the mMESSAGE mMACHINE T7 kit (Thermo Fisher Scientific). .. The DNA template for in vitro transcription containing the humanized Streptococcus pyogenes Cas9 gene was PCR-amplified from the px330 plasmid. eTw5-7 deletion mice were generated by injecting a mix of Cas9 mRNA (final concentration of 100 ng/ul) and sgRNA (50 ng/ul) in injection buffer (10 mM Tris, pH 7.5; 0.1 mM EDTA) into the cytoplasm of C57black embryos in accordance with standard procedure approved by Ben-Gurion University.

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: For in vivo rescue experiments, Tmem63c cDNA was linearized using XbaI FD (Thermo Fisher Scientific) and purified using GeneJET Gel Extraction Kit (Thermo Fisher Scientific). .. In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion). .. For poly-A-tailing, the Poly(A)-tailing Kit (Ambion) was used, followed by RNA extraction by RNeasy Mini Kit (Qiagen).

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: DENV was produced by transfection of viral RNA into Huh7 cells and harvesting the culture supernatants at days 5–7. .. A Renilla reporter DENV2 NGC strain RNA was transcribed in vitro by mMessage/mMachine T7 kit (Ambion) from pACYC-Rluc2A-NGC linearized by XbaI. .. DENV was produced by electroporation of the viral RNA into BHK-21 cells and harvesting the supernatants at day 10.

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: Alanine-coding mutations for conserved and control residues were introduced into full-length cDNA clone pEAV211 ( ) using appropriate shuttle vectors and restriction enzymes. .. The presence of the mutations was confirmed by sequencing. pEAV plasmid DNA was in vitro transcribed with the mMessage-mMachine T7 kit (Ambion), and the synthesized RNA was transfected into BHK-21 cells after LiCl precipitation as described previously ( ). .. Virus replication was monitored by immunofluorescence microscopy until 72 h post transfection (p.t.) using antibodies directed against nsp3 and N protein as described ( ) and by plaque assays ( ) using transfected cell culture supernatants, to monitor the production of viral progeny.

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: Open reading frames (ORFs) encoding 26 hits were selected from the Human ORFeome library of cDNA clones( ) (Open Biosystems), three from Addgene and one from DNASU( ) and recombined into a pFLAG (for FLAG tagging) vector using Gateway technology (Invitrogen). .. DENV2 16681 strain RNA was transcribed in vitro using mMessage/mMachine T7 kit (Ambion) from pD2IC-30P-NBX plasmid linearized by XbaI. .. DENV was produced by transfection of viral RNA into Huh7 cells and harvesting the culture supernatants at days 5–7.

    Quantitation Assay:

    Article Title: Recombinant Porcine Reproductive and Respiratory Syndrome Virus Expressing Membrane-Bound Interleukin-15 as an Immunomodulatory Adjuvant Enhances NK and γδ T Cell Responses and Confers Heterologous Protection
    Article Snippet: Paragraph title: Quantitation of viral RNA loads in sera and lung tissues. ... The RNA standard used for qRT-PCR was derived from the in vitro transcription of a PRRSV full-length cDNA clone, pACYC-VR2385, by using the mMessage mMachine T7 kit (Ambion).

    Produced:

    Article Title: Targeting fidelity of adenine and cytosine base editors in mouse embryos
    Article Snippet: The sgRNAs were designed using CRISPR.MIT.EDU, and subsequently produced using ThermoFisher’s sgRNA In Vitro Transcription Service. .. ABE, BE4, and VQR-BE3 mRNAs were synthesized in vitro using the mMESSAGE mMACHINE T7 kit (ThermoFisher Scientific).

    Article Title: An allosteric link connecting the lipid-protein interface to the gating of the nicotinic acetylcholine receptor
    Article Snippet: Each nAChR subunit DNA was transferred into the pcDNA3 vector as an EcoRI fragment. .. The α1 , β1 , δ, and ε nAChR-pcDNA3 was linearized with XhoI and capped cRNA was produced by in vitro transcription using the mMESSAGE mMACHINE® T7 kit (Ambion). .. All mutants were created using QuikChange™ Site-Directed Mutagenesis kits (Agilent) and verified by sequencing.

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: DENV was produced by transfection of viral RNA into Huh7 cells and harvesting the culture supernatants at days 5–7. .. A Renilla reporter DENV2 NGC strain RNA was transcribed in vitro by mMessage/mMachine T7 kit (Ambion) from pACYC-Rluc2A-NGC linearized by XbaI.

    Article Title: Single-cell transcriptional dynamics of flavivirus infection
    Article Snippet: DENV2 16681 strain RNA was transcribed in vitro using mMessage/mMachine T7 kit (Ambion) from pD2IC-30P-NBX plasmid linearized by XbaI. .. DENV2 16681 strain RNA was transcribed in vitro using mMessage/mMachine T7 kit (Ambion) from pD2IC-30P-NBX plasmid linearized by XbaI.

    Concentration Assay:

    Article Title: A missense mutation in ASRGL1 is involved in causing autosomal recessive retinal degeneration
    Article Snippet: The recombinant wt-h ASRGL1 and G178R -h ASRGL1 clones were used to generate the full-length mRNA with the mMESSAGE mMACHINE T7 kit (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions. .. Each embryo was injected with a specific dose of mRNA (ranging from 1 to 200 pg) of wt-h ASRGL1 or G178R-h ASRGL1 mRNA and/or phenol red dye.

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion). .. In vitro transcription of capped RNA and following TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    Article Title: Analysis of the genomic architecture of a complex trait locus in hypertensive rat models links Tmem63c to kidney damage
    Article Snippet: In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion). .. In vitro transcription of capped RNA followed by TurboDNase treatment were performed using mMessage mMachine T7 Kit (Ambion).

    BAC Assay:

    Article Title: Discovery of an essential nucleotidylating activity associated with a newly delineated conserved domain in the RNA polymerase-containing protein of all nidoviruses
    Article Snippet: Mutations in the SARS-CoV nsp12-coding region were engineered in prSCV, a pBeloBac11 derivative containing a full-length cDNA copy of the SARS-CoV Frankfurt-1 sequence ( ) by using ‘en passant recombineering’ as described in Tischer et al . ( ). .. The (mutated) BAC DNA was linearized with NotI, extracted with phenol–chloroform, and transcribed with T7 RNA Polymerase (mMessage-mMachine T7 kit; Ambion) using an input of 2 μg of BAC DNA per 20-μl reaction. .. Viral RNA transcripts were precipitated with LiCl according to the manufacturer's protocol.

    Staining:

    Article Title: Far Upstream Element-Binding Protein 1 Binds the 3′ Untranslated Region of PKD2 and Suppresses Its Translation
    Article Snippet: Capped human fubp1 mRNA was synthesized using the mMessage mMachine T7 kit (Ambion, Austin, TX) and injected into fertilized embryos (at 200 pg each). .. Capped human fubp1 mRNA was synthesized using the mMessage mMachine T7 kit (Ambion, Austin, TX) and injected into fertilized embryos (at 200 pg each).

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Thermo Fisher mmessage mmachine t7 transcription kit
    Mmessage Mmachine T7 Transcription Kit, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 204 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/mmessage mmachine t7 transcription kit/product/Thermo Fisher
    Average 99 stars, based on 204 article reviews
    Price from $9.99 to $1999.99
    mmessage mmachine t7 transcription kit - by Bioz Stars, 2019-10
    99/100 stars
      Buy from Supplier

    Image Search Results