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InGex tgirt iii
Tgirt Iii, supplied by InGex, used in various techniques. Bioz Stars score: 97/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/tgirt iii/product/InGex
Average 97 stars, based on 2 article reviews
Price from $9.99 to $1999.99
tgirt iii - by Bioz Stars, 2020-04
97/100 stars

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Related Articles

Clone Assay:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: Briefly, the desired HDR donor was first cloned downstream of a T7 promoter and the T7-HDR donor sequence amplified by PCR. .. RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex).

Amplification:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: Briefly, the desired HDR donor was first cloned downstream of a T7 promoter and the T7-HDR donor sequence amplified by PCR. .. RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex).

Article Title: TUT‐DIS3L2 is a mammalian surveillance pathway for aberrant structured non‐coding RNAs
Article Snippet: .. PCR amplification of snRNAs After RNA immunoprecipitation, the isolated RNA was ligated with L3AppDNA linker (as in 5′ UTR PCR) and reverse‐transcribed for 50 min. at 60°C using TGIRT‐III (InGex) and RT‐CLIP2 primer following manufacturer's instructions. .. PCR was performed with RT‐CLIP2 and indicated gene‐specific primers ( ).

Synthesized:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: .. RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex). ..

Neutralization:

Article Title: Template-switching mechanism of a group II intron-encoded reverse transcriptase and its implications for biological function and RNA-Seq
Article Snippet: RNA-Seq libraries were prepared as described previously ( , ) by using TGIRT-III (InGex), a commercial version of GsI–IIC RT, with different R2 RNA/R2R starter duplexes and acceptor nucleic acids as indicated in the text. .. RNA-Seq libraries were prepared as described previously ( , ) by using TGIRT-III (InGex), a commercial version of GsI–IIC RT, with different R2 RNA/R2R starter duplexes and acceptor nucleic acids as indicated in the text.

Electrophoresis:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex). .. The reaction was quenched with HCl, the final ssDNA product purified using Ampure XP magnetic beads (Beckman Coulter) and eluted in sterile RNAse-free H2O. ssDNA quality was analysed by capillary electrophoresis (Bioanalyzer, Agilent).

Incubation:

Article Title: DMS-MaPseq for genome-wide or targeted RNA structure probing in vivo
Article Snippet: MgCl2 was added to a 10 mM final concentration, and 3 μl of Hybridase Thermostable RNase H (Epicentre) was added, followed by a 30 min incubation at 45°C. .. 20–100 ng of RNA was used for reverse transcription with 100 U TGIRT-III (InGex) for 2h at 57°C in the same TGIRT reaction conditions described above.

Article Title: LC/MS analysis and deep sequencing reveal the accurate RNA composition in the HIV-1 virion
Article Snippet: Reverse transcription with TGIRT™-III (InGex, 1 µL, 500 nM) was performed in 19 µL of reaction buffer (450 mM NaCl, 5 mM MgCl2 , 20 mM Tris-HCl, pH 7.5), with DTT (5 mM) and RT primer (5 µM) for 30 min at room temperature. .. After 30 min dNTPs (1.25 mM each, final volume 20 µL) were added and the reaction was incubated at 60 °C for 50 min.

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex). .. Reactions were then incubated at 60°C for 15 min and terminated by adding 5 N NaOH to a final concentration of 0.25 N and incubated at 95°C for 3 min to degrade RNAs and denature protein.

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: Reverse transcription with TGIRT-III (InGex) was initiated from a DNA primer (5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTTN-3') encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself. .. Reactions were incubated at 60°C for 15 min and were terminated by adding 5 N NaOH to a final concentration of 0.25 N and incubated at 95°C for 3 min to degrade RNAs and denature protein.

Modification:

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: TGIRT-seq libraries were prepared essentially as described , with a modification in the starting molecule (see below). .. Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex).

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: TGIRT-seq libraries TGIRT-seq libraries were prepared with a modification of the Total RNA-seq method [ ]. .. Reverse transcription with TGIRT-III (InGex) was initiated from a DNA primer (5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTTN-3') encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself.

Hybridization:

Article Title: DMS-MaPseq for genome-wide or targeted RNA structure probing in vivo
Article Snippet: RNase H subtraction was performed by adding 5 μg of published subtraction oligos in a total volume of 30 μl in 1X Hybridization Buffer (200 mM NaCl, 100 mM Tris pH 7.5). .. 20–100 ng of RNA was used for reverse transcription with 100 U TGIRT-III (InGex) for 2h at 57°C in the same TGIRT reaction conditions described above.

Ligation:

Article Title: LC/MS analysis and deep sequencing reveal the accurate RNA composition in the HIV-1 virion
Article Snippet: Ligation at the 3′-end of RNA was performed at 4 °C for 72 h in dephosphorylation buffer, containing in addition, 15% DMSO, 5 µM adenylated 3′-RNA adaptor, 0.5 U/µL T4 RNA ligase (ThermoFisher Scientific) and 1 U/µL T4 RNA ligase 2 truncated (NEB). .. Reverse transcription with TGIRT™-III (InGex, 1 µL, 500 nM) was performed in 19 µL of reaction buffer (450 mM NaCl, 5 mM MgCl2 , 20 mM Tris-HCl, pH 7.5), with DTT (5 mM) and RT primer (5 µM) for 30 min at room temperature.

other:

Article Title: Monitored eCLIP: high accuracy mapping of RNA-protein interactions
Article Snippet: As this ability may differ from one RTase to another, we repeated eIF4A3-HA meCLIP with three additional RTases: AffinityScript, SuperScript III and TGIRT III .

Article Title: Monitored eCLIP: high accuracy mapping of RNA-protein interactions
Article Snippet: TGIRT-III (InGex) was used at 60°C, as previously described ( ).

Article Title: Monitored eCLIP: high accuracy mapping of RNA-protein interactions
Article Snippet: As this ability may differ from one RTase to another, we repeated eIF4A3-HA meCLIP with three additional RTases: AffinityScript, SuperScript III and TGIRT III .

Sequencing:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: Briefly, the desired HDR donor was first cloned downstream of a T7 promoter and the T7-HDR donor sequence amplified by PCR. .. RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex).

Article Title: LC/MS analysis and deep sequencing reveal the accurate RNA composition in the HIV-1 virion
Article Snippet: Paragraph title: Deep sequencing library preparation ... Reverse transcription with TGIRT™-III (InGex, 1 µL, 500 nM) was performed in 19 µL of reaction buffer (450 mM NaCl, 5 mM MgCl2 , 20 mM Tris-HCl, pH 7.5), with DTT (5 mM) and RT primer (5 µM) for 30 min at room temperature.

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex). .. Reverse transcription by TGIRT-III is initiated by template switching from a starting molecule consisting of a DNA primer ( 5’-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTATTAN-3’ ) encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself.

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: .. Reverse transcription with TGIRT-III (InGex) was initiated from a DNA primer (5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTTN-3') encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself. .. Reactions contained purified RNAs, reaction medium (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting annealed molecule and 1 μM TGIRT-III.

Binding Assay:

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex). .. Reverse transcription by TGIRT-III is initiated by template switching from a starting molecule consisting of a DNA primer ( 5’-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTATTAN-3’ ) encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself.

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: .. Reverse transcription with TGIRT-III (InGex) was initiated from a DNA primer (5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTTN-3') encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself. .. Reactions contained purified RNAs, reaction medium (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting annealed molecule and 1 μM TGIRT-III.

RNA Sequencing Assay:

Article Title: Template-switching mechanism of a group II intron-encoded reverse transcriptase and its implications for biological function and RNA-Seq
Article Snippet: .. RNA-Seq libraries were prepared as described previously ( , ) by using TGIRT-III (InGex), a commercial version of GsI–IIC RT, with different R2 RNA/R2R starter duplexes and acceptor nucleic acids as indicated in the text. .. After terminating the reactions with NaOH and neutralization with HCl as described above for template-switching reactions, the volume was raised to 100 μl with H2 O, and cDNA products containing the R2R adapter attached to their 5′ end were cleaned-up by using a MinElute column (Qiagen) to remove unused primer.

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: TGIRT-seq libraries TGIRT-seq libraries were prepared with a modification of the Total RNA-seq method [ ]. .. Reverse transcription with TGIRT-III (InGex) was initiated from a DNA primer (5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTTN-3') encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself.

Magnetic Beads:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex). .. The reaction was quenched with HCl, the final ssDNA product purified using Ampure XP magnetic beads (Beckman Coulter) and eluted in sterile RNAse-free H2O. ssDNA quality was analysed by capillary electrophoresis (Bioanalyzer, Agilent).

Isolation:

Article Title: TUT‐DIS3L2 is a mammalian surveillance pathway for aberrant structured non‐coding RNAs
Article Snippet: .. PCR amplification of snRNAs After RNA immunoprecipitation, the isolated RNA was ligated with L3AppDNA linker (as in 5′ UTR PCR) and reverse‐transcribed for 50 min. at 60°C using TGIRT‐III (InGex) and RT‐CLIP2 primer following manufacturer's instructions. .. PCR was performed with RT‐CLIP2 and indicated gene‐specific primers ( ).

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: RNA extraction and TGIRT-seq library preparation Cellular and EV RNAs were extracted by using a mirVana miRNA Isolation Kit. .. Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex).

Purification:

Article Title: DMS-MaPseq for genome-wide or targeted RNA structure probing in vivo
Article Snippet: The reaction was again purified by Zymo RNA Clean & Concentrator-5 column to deplete small RNA species, followed by treatment with DNaseI (Ambion) per manufacturer instructions and a final column clean-up to remove excess RNase H subtraction oligos. .. 20–100 ng of RNA was used for reverse transcription with 100 U TGIRT-III (InGex) for 2h at 57°C in the same TGIRT reaction conditions described above.

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex). .. The reaction was quenched with HCl, the final ssDNA product purified using Ampure XP magnetic beads (Beckman Coulter) and eluted in sterile RNAse-free H2O. ssDNA quality was analysed by capillary electrophoresis (Bioanalyzer, Agilent).

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: .. Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex). .. Reverse transcription by TGIRT-III is initiated by template switching from a starting molecule consisting of a DNA primer ( 5’-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTATTAN-3’ ) encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself.

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: .. Reactions contained purified RNAs, reaction medium (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting annealed molecule and 1 μM TGIRT-III. ..

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: Reverse transcription with TGIRT-III (InGex) was initiated from a DNA primer (5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTTN-3') encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself. .. Reactions contained purified RNAs, reaction medium (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting annealed molecule and 1 μM TGIRT-III.

Polymerase Chain Reaction:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: Briefly, the desired HDR donor was first cloned downstream of a T7 promoter and the T7-HDR donor sequence amplified by PCR. .. RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex).

Article Title: TUT‐DIS3L2 is a mammalian surveillance pathway for aberrant structured non‐coding RNAs
Article Snippet: .. PCR amplification of snRNAs After RNA immunoprecipitation, the isolated RNA was ligated with L3AppDNA linker (as in 5′ UTR PCR) and reverse‐transcribed for 50 min. at 60°C using TGIRT‐III (InGex) and RT‐CLIP2 primer following manufacturer's instructions. .. PCR was performed with RT‐CLIP2 and indicated gene‐specific primers ( ).

De-Phosphorylation Assay:

Article Title: LC/MS analysis and deep sequencing reveal the accurate RNA composition in the HIV-1 virion
Article Snippet: Ligation at the 3′-end of RNA was performed at 4 °C for 72 h in dephosphorylation buffer, containing in addition, 15% DMSO, 5 µM adenylated 3′-RNA adaptor, 0.5 U/µL T4 RNA ligase (ThermoFisher Scientific) and 1 U/µL T4 RNA ligase 2 truncated (NEB). .. Reverse transcription with TGIRT™-III (InGex, 1 µL, 500 nM) was performed in 19 µL of reaction buffer (450 mM NaCl, 5 mM MgCl2 , 20 mM Tris-HCl, pH 7.5), with DTT (5 mM) and RT primer (5 µM) for 30 min at room temperature.

Concentration Assay:

Article Title: DMS-MaPseq for genome-wide or targeted RNA structure probing in vivo
Article Snippet: MgCl2 was added to a 10 mM final concentration, and 3 μl of Hybridase Thermostable RNase H (Epicentre) was added, followed by a 30 min incubation at 45°C. .. 20–100 ng of RNA was used for reverse transcription with 100 U TGIRT-III (InGex) for 2h at 57°C in the same TGIRT reaction conditions described above.

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex). .. Reactions were then incubated at 60°C for 15 min and terminated by adding 5 N NaOH to a final concentration of 0.25 N and incubated at 95°C for 3 min to degrade RNAs and denature protein.

Article Title: BCDIN3D regulates tRNAHis 3’ fragment processing
Article Snippet: Reverse transcription with TGIRT-III (InGex) was initiated from a DNA primer (5'-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTTN-3') encoding the reverse complement of the Illumina Read2 sequencing primer binding site (R2R) annealed to a complementary RNA oligonucleotide (R2) such that there is a single nucleotide 3’ DNA overhang composed of an equimolar mixture of A, G, C and T. The RNA oligonucleotide is blocked at its 3’ end with C3Sp (IDT) to inhibit template switching to itself. .. Reactions were incubated at 60°C for 15 min and were terminated by adding 5 N NaOH to a final concentration of 0.25 N and incubated at 95°C for 3 min to degrade RNAs and denature protein.

RNA Extraction:

Article Title: Distinct mechanisms of microRNA sorting into cancer cell-derived extracellular vesicle subtypes
Article Snippet: Paragraph title: RNA extraction and TGIRT-seq library preparation ... Reverse transcription reactions contained purified RNAs, buffer (20 mM Tris-HCl, pH7.5, 450 mM NaCl, 5 mM MgCl2 ), 5 mM DTT, 100 nM starting molecule (see below) and 1 µM TGIRT-III (Ingex).

In Vitro:

Article Title: Reprogramming human T cell function and specificity with non-viral genome targeting
Article Snippet: .. RNA was synthesized by in vitro transcription using HiScribe T7 RNA polymerase (New England Biolabs) and reverse-transcribed using TGIRT-III (InGex). ..

Immunoprecipitation:

Article Title: TUT‐DIS3L2 is a mammalian surveillance pathway for aberrant structured non‐coding RNAs
Article Snippet: .. PCR amplification of snRNAs After RNA immunoprecipitation, the isolated RNA was ligated with L3AppDNA linker (as in 5′ UTR PCR) and reverse‐transcribed for 50 min. at 60°C using TGIRT‐III (InGex) and RT‐CLIP2 primer following manufacturer's instructions. .. PCR was performed with RT‐CLIP2 and indicated gene‐specific primers ( ).

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    InGex tgirt iii rt
    <t>TGIRT-seq</t> reads map mostly to protein-coding genes but with greater representation of small ncRNAs than TruSeq libraries. ( A ) for different library preparation methods for numbered replicates of Samples A–D. ( B ) Stacked bar graphs showing the percentage of small noncoding RNA reads that map to different classes of small ncRNAs for different library preparation methods for numbered replicates of Samples A–D. MiscRNA includes ribozymes, such as RNase P RNA, imprinted transcripts, such as Xist, and other transcripts that cannot be classified into other RNA annotation categories. ( Left panels) TGIRT-seq; ( middle panels) TruSeq v2 (from ABRF at <t>three</t> different sites, L/R/V); ( right panels) TruSeq v3 (from ABRF at site W). Features and small ncRNA classes are color coded as indicated to the right of the bar graphs.
    Tgirt Iii Rt, supplied by InGex, used in various techniques. Bioz Stars score: 89/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/tgirt iii rt/product/InGex
    Average 89 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    tgirt iii rt - by Bioz Stars, 2020-04
    89/100 stars
      Buy from Supplier

    93
    InGex recombinant tgirt iii
    ER-RNA expression in transgenic mouse models of DM1. ( A ) cDNA slot blot of total cellular RNA from HSA LR quadriceps (Quads, 2 μg, three different mice) using <t>Superscript-III</t> (SS-III) or <t>TGIRT-III.</t> ( B ) cDNA slot blot of 2 μg total cellular RNA from quadriceps or tibialis anterior (TA) muscles from HSA LR and HSA XLR mice. Bottom-most wells in each column are from HSA SR or HSA NR mice, which do not express ER-RNA. ( C ) Relative amount of ER-RNA by cDNA slot blot in HSA LR and HSA XLR quadriceps and TA muscle (black bars, mean signal in HSA LR quadriceps set to 1), as compared to inferred ER-RNA level based on qRT-PCR (white bars, calculated by multiplying expression level x repeat length, with the mean product in HSA LR quadriceps set to 1). Results are based on n = 3 for HSA LR and HSA XLR mice. HSA transgene expression by qRT-PCR was normalized to general transcription factor 2b (Gtf2b) .
    Recombinant Tgirt Iii, supplied by InGex, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/recombinant tgirt iii/product/InGex
    Average 93 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    recombinant tgirt iii - by Bioz Stars, 2020-04
    93/100 stars
      Buy from Supplier

    95
    InGex tgirt iii enzyme
    <t>TGIRT</t> ssDNA-seq workflow. The target DNA (2–50 ng in this work) is treated with alkaline phosphatase to remove 3′ phosphates and heat denatured prior to DNA-seq library construction. The resulting ssDNAs with 3′ OH termini are then used for TGIRT template-switching DNA synthesis coupled to Illumina read 2 reverse (R2R) DNA-seq adapter addition. In this novel reaction, the <t>TGIRT-III</t> enzyme (InGex) binds first to a synthetic 34-bp R2 RNA/R2R DNA heteroduplex in which the R2R DNA primer has a single nucleotide 3′ overhang that can direct TGIRT template switching by base pairing to the 3′ nucleotide of a target DNA strand. For minimally biased library preparation, the 3′-overhang nucleotide is an equimolar mixture of A, C, G, and T (denoted N). A 3′-blocking group (C3 spacer; 3′SpC3) is attached to the end of the R2 RNA oligonucleotide to prevent template-switching to that RNA. After the TGIRT enzyme extends the DNA primer to produce a DNA copy of the target DNA strand with an R2R adapter seamlessly linked to its 5′ end, a 5′ adenylated (App) Illumina read 1 reverse (R1R) adapter is added to its 3′ end by single-stranded DNA ligation using a Thermostable 5′ AppDNA/RNA ligase (New England Biolabs). In the workflow shown, an unique molecular identifier (UMI) is positioned at the 5′ end of the R1R DNA oligonucleotide. A final PCR step adds flow cell capture sites and barcodes for Illumina sequencing and sample multiplexing.
    Tgirt Iii Enzyme, supplied by InGex, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/tgirt iii enzyme/product/InGex
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    tgirt iii enzyme - by Bioz Stars, 2020-04
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    Image Search Results


    TGIRT-seq reads map mostly to protein-coding genes but with greater representation of small ncRNAs than TruSeq libraries. ( A ) for different library preparation methods for numbered replicates of Samples A–D. ( B ) Stacked bar graphs showing the percentage of small noncoding RNA reads that map to different classes of small ncRNAs for different library preparation methods for numbered replicates of Samples A–D. MiscRNA includes ribozymes, such as RNase P RNA, imprinted transcripts, such as Xist, and other transcripts that cannot be classified into other RNA annotation categories. ( Left panels) TGIRT-seq; ( middle panels) TruSeq v2 (from ABRF at three different sites, L/R/V); ( right panels) TruSeq v3 (from ABRF at site W). Features and small ncRNA classes are color coded as indicated to the right of the bar graphs.

    Journal: RNA

    Article Title: RNA-seq of human reference RNA samples using a thermostable group II intron reverse transcriptase

    doi: 10.1261/rna.055558.115

    Figure Lengend Snippet: TGIRT-seq reads map mostly to protein-coding genes but with greater representation of small ncRNAs than TruSeq libraries. ( A ) for different library preparation methods for numbered replicates of Samples A–D. ( B ) Stacked bar graphs showing the percentage of small noncoding RNA reads that map to different classes of small ncRNAs for different library preparation methods for numbered replicates of Samples A–D. MiscRNA includes ribozymes, such as RNase P RNA, imprinted transcripts, such as Xist, and other transcripts that cannot be classified into other RNA annotation categories. ( Left panels) TGIRT-seq; ( middle panels) TruSeq v2 (from ABRF at three different sites, L/R/V); ( right panels) TruSeq v3 (from ABRF at site W). Features and small ncRNA classes are color coded as indicated to the right of the bar graphs.

    Article Snippet: Half of the recovered RNA was used for cDNA synthesis via TGIRT template switching with 1 μM TGIRT-III RT (InGex, LLC) for 15 min at 60°C, as previously described ( ).

    Techniques:

    ER-RNA expression in transgenic mouse models of DM1. ( A ) cDNA slot blot of total cellular RNA from HSA LR quadriceps (Quads, 2 μg, three different mice) using Superscript-III (SS-III) or TGIRT-III. ( B ) cDNA slot blot of 2 μg total cellular RNA from quadriceps or tibialis anterior (TA) muscles from HSA LR and HSA XLR mice. Bottom-most wells in each column are from HSA SR or HSA NR mice, which do not express ER-RNA. ( C ) Relative amount of ER-RNA by cDNA slot blot in HSA LR and HSA XLR quadriceps and TA muscle (black bars, mean signal in HSA LR quadriceps set to 1), as compared to inferred ER-RNA level based on qRT-PCR (white bars, calculated by multiplying expression level x repeat length, with the mean product in HSA LR quadriceps set to 1). Results are based on n = 3 for HSA LR and HSA XLR mice. HSA transgene expression by qRT-PCR was normalized to general transcription factor 2b (Gtf2b) .

    Journal: Nucleic Acids Research

    Article Title: Detection of expanded RNA repeats using thermostable group II intron reverse transcriptase

    doi: 10.1093/nar/gkx867

    Figure Lengend Snippet: ER-RNA expression in transgenic mouse models of DM1. ( A ) cDNA slot blot of total cellular RNA from HSA LR quadriceps (Quads, 2 μg, three different mice) using Superscript-III (SS-III) or TGIRT-III. ( B ) cDNA slot blot of 2 μg total cellular RNA from quadriceps or tibialis anterior (TA) muscles from HSA LR and HSA XLR mice. Bottom-most wells in each column are from HSA SR or HSA NR mice, which do not express ER-RNA. ( C ) Relative amount of ER-RNA by cDNA slot blot in HSA LR and HSA XLR quadriceps and TA muscle (black bars, mean signal in HSA LR quadriceps set to 1), as compared to inferred ER-RNA level based on qRT-PCR (white bars, calculated by multiplying expression level x repeat length, with the mean product in HSA LR quadriceps set to 1). Results are based on n = 3 for HSA LR and HSA XLR mice. HSA transgene expression by qRT-PCR was normalized to general transcription factor 2b (Gtf2b) .

    Article Snippet: Purified recombinant TGIRT-III was prepared as previously described ( ) or obtained from Ingex, St. Louis, MO, USA.

    Techniques: RNA Expression, Transgenic Assay, Dot Blot, Mouse Assay, Quantitative RT-PCR, Expressing

    Reverse transcription across ER-RNA tracts using TGIRT-III or SS-III. ( A ) Diagram of ER-RNA showing HEX-labeled primer annealed 3′ of repeat tract and unlabeled primers tiled across the expanded repeat. ( B ) Representative chromatogram of cDNA generated by TGIRT-III from r(CUG) 100 in the presence or absence of intervening repeat primers. ( C ) Representative chromatogram of cDNA generated by TGIRT-III from r(GGGGCC) 40 , as in (B). ( D and E ) Representative chromatograms from r(CUG) 100 and r(GGGGCC) 40 templates using Superscript-III, as in (B) and (C).

    Journal: Nucleic Acids Research

    Article Title: Detection of expanded RNA repeats using thermostable group II intron reverse transcriptase

    doi: 10.1093/nar/gkx867

    Figure Lengend Snippet: Reverse transcription across ER-RNA tracts using TGIRT-III or SS-III. ( A ) Diagram of ER-RNA showing HEX-labeled primer annealed 3′ of repeat tract and unlabeled primers tiled across the expanded repeat. ( B ) Representative chromatogram of cDNA generated by TGIRT-III from r(CUG) 100 in the presence or absence of intervening repeat primers. ( C ) Representative chromatogram of cDNA generated by TGIRT-III from r(GGGGCC) 40 , as in (B). ( D and E ) Representative chromatograms from r(CUG) 100 and r(GGGGCC) 40 templates using Superscript-III, as in (B) and (C).

    Article Snippet: Purified recombinant TGIRT-III was prepared as previously described ( ) or obtained from Ingex, St. Louis, MO, USA.

    Techniques: Labeling, Generated

    Reverse transcription of enzymatically synthesized ER-RNAs. ( A ) Diagram of ER-RNA template, multiply primed with 5′ HEX end-labeled oligonucleotides. Representative chromatograms of HEX-labeled cDNA generated by TGIRT-III ( B–E ) or Superscript-III ( F–I ). Each trace represents analysis of a single cDNA synthesis.

    Journal: Nucleic Acids Research

    Article Title: Detection of expanded RNA repeats using thermostable group II intron reverse transcriptase

    doi: 10.1093/nar/gkx867

    Figure Lengend Snippet: Reverse transcription of enzymatically synthesized ER-RNAs. ( A ) Diagram of ER-RNA template, multiply primed with 5′ HEX end-labeled oligonucleotides. Representative chromatograms of HEX-labeled cDNA generated by TGIRT-III ( B–E ) or Superscript-III ( F–I ). Each trace represents analysis of a single cDNA synthesis.

    Article Snippet: Purified recombinant TGIRT-III was prepared as previously described ( ) or obtained from Ingex, St. Louis, MO, USA.

    Techniques: Synthesized, Labeling, Generated

    TGIRT ssDNA-seq workflow. The target DNA (2–50 ng in this work) is treated with alkaline phosphatase to remove 3′ phosphates and heat denatured prior to DNA-seq library construction. The resulting ssDNAs with 3′ OH termini are then used for TGIRT template-switching DNA synthesis coupled to Illumina read 2 reverse (R2R) DNA-seq adapter addition. In this novel reaction, the TGIRT-III enzyme (InGex) binds first to a synthetic 34-bp R2 RNA/R2R DNA heteroduplex in which the R2R DNA primer has a single nucleotide 3′ overhang that can direct TGIRT template switching by base pairing to the 3′ nucleotide of a target DNA strand. For minimally biased library preparation, the 3′-overhang nucleotide is an equimolar mixture of A, C, G, and T (denoted N). A 3′-blocking group (C3 spacer; 3′SpC3) is attached to the end of the R2 RNA oligonucleotide to prevent template-switching to that RNA. After the TGIRT enzyme extends the DNA primer to produce a DNA copy of the target DNA strand with an R2R adapter seamlessly linked to its 5′ end, a 5′ adenylated (App) Illumina read 1 reverse (R1R) adapter is added to its 3′ end by single-stranded DNA ligation using a Thermostable 5′ AppDNA/RNA ligase (New England Biolabs). In the workflow shown, an unique molecular identifier (UMI) is positioned at the 5′ end of the R1R DNA oligonucleotide. A final PCR step adds flow cell capture sites and barcodes for Illumina sequencing and sample multiplexing.

    Journal: Scientific Reports

    Article Title: Facile single-stranded DNA sequencing of human plasma DNA via thermostable group II intron reverse transcriptase template switching

    doi: 10.1038/s41598-017-09064-w

    Figure Lengend Snippet: TGIRT ssDNA-seq workflow. The target DNA (2–50 ng in this work) is treated with alkaline phosphatase to remove 3′ phosphates and heat denatured prior to DNA-seq library construction. The resulting ssDNAs with 3′ OH termini are then used for TGIRT template-switching DNA synthesis coupled to Illumina read 2 reverse (R2R) DNA-seq adapter addition. In this novel reaction, the TGIRT-III enzyme (InGex) binds first to a synthetic 34-bp R2 RNA/R2R DNA heteroduplex in which the R2R DNA primer has a single nucleotide 3′ overhang that can direct TGIRT template switching by base pairing to the 3′ nucleotide of a target DNA strand. For minimally biased library preparation, the 3′-overhang nucleotide is an equimolar mixture of A, C, G, and T (denoted N). A 3′-blocking group (C3 spacer; 3′SpC3) is attached to the end of the R2 RNA oligonucleotide to prevent template-switching to that RNA. After the TGIRT enzyme extends the DNA primer to produce a DNA copy of the target DNA strand with an R2R adapter seamlessly linked to its 5′ end, a 5′ adenylated (App) Illumina read 1 reverse (R1R) adapter is added to its 3′ end by single-stranded DNA ligation using a Thermostable 5′ AppDNA/RNA ligase (New England Biolabs). In the workflow shown, an unique molecular identifier (UMI) is positioned at the 5′ end of the R1R DNA oligonucleotide. A final PCR step adds flow cell capture sites and barcodes for Illumina sequencing and sample multiplexing.

    Article Snippet: Reactions were done with 5–50 ng DNA substrate, 100 nM annealed template-primer substrate, and TGIRT-III enzyme (400 units, 1 mM; InGex, LLC; St. Louis) in 20 μl of reaction medium containing 420 mM NaCl, 5 mM MgCl2 , 20 mM Tris-HCl, pH 7.5, 5 mM DTT and 1 mM dNTPs (an equimolar mix of 1 mM dATP, dCTP, dGTP, and dTTP).

    Techniques: DNA Sequencing, DNA Synthesis, Blocking Assay, DNA Ligation, Polymerase Chain Reaction, Flow Cytometry, Sequencing, Multiplexing

    Bisulfite sequencing of plasma cfDNA. Three separate TGIRT-seq libraries were each constructed from ~5 ng of bisulfite-treated plasma DNA from a healthy male individual, and sequenced to obtain TGIRT-seq datasets BPD1-3 (see Supplementary Table S4 ). The datasets were combined and analyzed to identify DNA methylation sites, as described in Methods. ( a ) Annotation of DNA methylation sites in the human genome and determination of DNA methylation densities by TGIRT-seq of bisulfite-treated plasma cfDNA. Tracks from the inner to the outer circle represent: (1) annotations of Type II biomarkers that are highly variable in methylation density across tissues; (2) annotations of Type I biomarkers that are highly tissue specific (color-coded); (3) bar graphs of methylation densities within the annotated regions based on TGIRT-seq of bisulfite-treated plasma cfDNA; (4) genome coordinates. ( b ) Tissues-of-origin of plasma cfDNA from a healthy male individual determined by TGIRT-seq of bisulfite-treated DNA. The pie chart shows the percent contributions of different tissues in combined datasets PB1-3 determined by quadratic programming 7 . Neutrophils are derived from myeloid precursors, and lymphocytes are a combination of T-cells and B-cells. Comparison of tissues-of-origin of plasma cfDNA determined as in panel ( b ) from the three independent datasets BPD1-3 are shown in Supplementary Fig. 11 . Pearson’s correlation coefficients were ≥0.96 for each pairwise combination of the three individual datasets.

    Journal: Scientific Reports

    Article Title: Facile single-stranded DNA sequencing of human plasma DNA via thermostable group II intron reverse transcriptase template switching

    doi: 10.1038/s41598-017-09064-w

    Figure Lengend Snippet: Bisulfite sequencing of plasma cfDNA. Three separate TGIRT-seq libraries were each constructed from ~5 ng of bisulfite-treated plasma DNA from a healthy male individual, and sequenced to obtain TGIRT-seq datasets BPD1-3 (see Supplementary Table S4 ). The datasets were combined and analyzed to identify DNA methylation sites, as described in Methods. ( a ) Annotation of DNA methylation sites in the human genome and determination of DNA methylation densities by TGIRT-seq of bisulfite-treated plasma cfDNA. Tracks from the inner to the outer circle represent: (1) annotations of Type II biomarkers that are highly variable in methylation density across tissues; (2) annotations of Type I biomarkers that are highly tissue specific (color-coded); (3) bar graphs of methylation densities within the annotated regions based on TGIRT-seq of bisulfite-treated plasma cfDNA; (4) genome coordinates. ( b ) Tissues-of-origin of plasma cfDNA from a healthy male individual determined by TGIRT-seq of bisulfite-treated DNA. The pie chart shows the percent contributions of different tissues in combined datasets PB1-3 determined by quadratic programming 7 . Neutrophils are derived from myeloid precursors, and lymphocytes are a combination of T-cells and B-cells. Comparison of tissues-of-origin of plasma cfDNA determined as in panel ( b ) from the three independent datasets BPD1-3 are shown in Supplementary Fig. 11 . Pearson’s correlation coefficients were ≥0.96 for each pairwise combination of the three individual datasets.

    Article Snippet: Reactions were done with 5–50 ng DNA substrate, 100 nM annealed template-primer substrate, and TGIRT-III enzyme (400 units, 1 mM; InGex, LLC; St. Louis) in 20 μl of reaction medium containing 420 mM NaCl, 5 mM MgCl2 , 20 mM Tris-HCl, pH 7.5, 5 mM DTT and 1 mM dNTPs (an equimolar mix of 1 mM dATP, dCTP, dGTP, and dTTP).

    Techniques: Methylation Sequencing, Construct, DNA Methylation Assay, Methylation, Derivative Assay

    Testing reverse transcriptase conditions with eCLIP (A) eCLIP overall schematic. A single biological sample was lysed, immunoprecipitated, and taken through standard eCLIP library preparation until the reverse transcription stage, at which point it was split into multiple conditions. (B) Immunoprecipitation (IP) western blot images for (left) TARDBP and (right) RBFOX2 eCLIP performed in HEK293XT cells. (C) Library yield obtained in eCLIP experiments for RBFOX2 (filled circles) and TARDBP (empty circles), normalized to a Superscript III condition performed within that experiment batch. Average across all experiments is indicated by red dashed lines. For eCLIP experiments that were completed and sequenced (black), yield was calculated as the number of PCR cycles required to obtain 100 femtomoles of library (extrapolated from the library yield and number of PCR cycles performed). For additional experiments only taken to pre-amplified library stage (blue), library yield was determined as the Ct value obtained by qPCR of the pre-amplified library with standard library amplification primers. Reverse transcription conditions tested were AffinityScript (AffSc), Superscript II (SS2), Superscript III (SS3), Superscript IV (SS4), Superscript IV in manganese buffer (SS4Mn), Superscript IV in Superscript III buffer (SS4in3B), TGIRT-III enzyme (TGIRT), AMV, and M-MLV. Standard buffers and reaction conditions were used unless otherwise indicated (see Materials Methods).

    Journal: Methods (San Diego, Calif.)

    Article Title: Variation in single-nucleotide sensitivity of eCLIP derived from reverse transcription conditions

    doi: 10.1016/j.ymeth.2017.08.002

    Figure Lengend Snippet: Testing reverse transcriptase conditions with eCLIP (A) eCLIP overall schematic. A single biological sample was lysed, immunoprecipitated, and taken through standard eCLIP library preparation until the reverse transcription stage, at which point it was split into multiple conditions. (B) Immunoprecipitation (IP) western blot images for (left) TARDBP and (right) RBFOX2 eCLIP performed in HEK293XT cells. (C) Library yield obtained in eCLIP experiments for RBFOX2 (filled circles) and TARDBP (empty circles), normalized to a Superscript III condition performed within that experiment batch. Average across all experiments is indicated by red dashed lines. For eCLIP experiments that were completed and sequenced (black), yield was calculated as the number of PCR cycles required to obtain 100 femtomoles of library (extrapolated from the library yield and number of PCR cycles performed). For additional experiments only taken to pre-amplified library stage (blue), library yield was determined as the Ct value obtained by qPCR of the pre-amplified library with standard library amplification primers. Reverse transcription conditions tested were AffinityScript (AffSc), Superscript II (SS2), Superscript III (SS3), Superscript IV (SS4), Superscript IV in manganese buffer (SS4Mn), Superscript IV in Superscript III buffer (SS4in3B), TGIRT-III enzyme (TGIRT), AMV, and M-MLV. Standard buffers and reaction conditions were used unless otherwise indicated (see Materials Methods).

    Article Snippet: To each was added 3 μL H2 O, 4 μL of 5X TGIRT buffer (2.25M NaCl, 25 mM MgCl, 100 mM TrisHCl pH 7.5), 1 μL 0.1M DTT, 0.5 μL TGIRT-III enzyme (InGex), and 2.5 μL 10 mM dNTP mix.

    Techniques: Immunoprecipitation, Western Blot, Polymerase Chain Reaction, Amplification, Real-time Polymerase Chain Reaction