low range rna molecular weight markers  (Thermo Fisher)


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    Structured Review

    Thermo Fisher low range rna molecular weight markers
    Genomic organization and expression strategy of Or-CD15 . ( A ) Screen shot of the RpL23A / Uhg7 region taken from the UCSC genome browser. The RpL23A-RA and Uhg7 transcripts are depicted in blue, the location of Or-CD15 and the newly identified transcript RpL23A-RB are shown in black. Below, sequence conservation as measured by phastcons and the coverage of a multiple genome alignment is indicated; ( B ) Northern blot analysis of adult fruit fly <t>RNA.</t> The genomic position of the utilised probe is outlined in ( A ). The band at about 100 nt corresponds to snoRNA Or-CD15 ; sizes are indicated by an RNA molecular weight ladder (RiboRuler Low Range, Life Technologies); ( C ) <t>PCR</t> experiments using primers annealing to RpL23A exon 1 and Uhg7 exon 4; amplification of the genomic DNA (gDNA) produces the expected fragment of about 2 kb; the same primers used in RT-PCR experiments successfully amplified a fragment of 1.2 kb, representative of the new RpL23A-RB transcript. In this transcript, RpL23A and Uhg7 sequences are fused to each other. On the right, a DNA molecular weight ladder (100bp DNA ladder, New England BioLabs, Ipswich, MA, USA) is given.
    Low Range Rna Molecular Weight Markers, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster"

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster

    Journal: Non-Coding RNA

    doi: 10.3390/ncrna1020139

    Genomic organization and expression strategy of Or-CD15 . ( A ) Screen shot of the RpL23A / Uhg7 region taken from the UCSC genome browser. The RpL23A-RA and Uhg7 transcripts are depicted in blue, the location of Or-CD15 and the newly identified transcript RpL23A-RB are shown in black. Below, sequence conservation as measured by phastcons and the coverage of a multiple genome alignment is indicated; ( B ) Northern blot analysis of adult fruit fly RNA. The genomic position of the utilised probe is outlined in ( A ). The band at about 100 nt corresponds to snoRNA Or-CD15 ; sizes are indicated by an RNA molecular weight ladder (RiboRuler Low Range, Life Technologies); ( C ) PCR experiments using primers annealing to RpL23A exon 1 and Uhg7 exon 4; amplification of the genomic DNA (gDNA) produces the expected fragment of about 2 kb; the same primers used in RT-PCR experiments successfully amplified a fragment of 1.2 kb, representative of the new RpL23A-RB transcript. In this transcript, RpL23A and Uhg7 sequences are fused to each other. On the right, a DNA molecular weight ladder (100bp DNA ladder, New England BioLabs, Ipswich, MA, USA) is given.
    Figure Legend Snippet: Genomic organization and expression strategy of Or-CD15 . ( A ) Screen shot of the RpL23A / Uhg7 region taken from the UCSC genome browser. The RpL23A-RA and Uhg7 transcripts are depicted in blue, the location of Or-CD15 and the newly identified transcript RpL23A-RB are shown in black. Below, sequence conservation as measured by phastcons and the coverage of a multiple genome alignment is indicated; ( B ) Northern blot analysis of adult fruit fly RNA. The genomic position of the utilised probe is outlined in ( A ). The band at about 100 nt corresponds to snoRNA Or-CD15 ; sizes are indicated by an RNA molecular weight ladder (RiboRuler Low Range, Life Technologies); ( C ) PCR experiments using primers annealing to RpL23A exon 1 and Uhg7 exon 4; amplification of the genomic DNA (gDNA) produces the expected fragment of about 2 kb; the same primers used in RT-PCR experiments successfully amplified a fragment of 1.2 kb, representative of the new RpL23A-RB transcript. In this transcript, RpL23A and Uhg7 sequences are fused to each other. On the right, a DNA molecular weight ladder (100bp DNA ladder, New England BioLabs, Ipswich, MA, USA) is given.

    Techniques Used: Expressing, Sequencing, Northern Blot, Molecular Weight, Polymerase Chain Reaction, Amplification, Reverse Transcription Polymerase Chain Reaction

    Related Articles

    DNA Extraction:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: DNA extraction, manipulation and labelling, RNA electrophoresis, and blotting were carried out according to [ ]. .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    SYBR Green Assay:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ]. .. All PCR reactions were carried out in a final volume of 15 μL using 1 μL of diluted cDNA, 7.5 μL of 2X SYBR-Green (Bio-Rad) and 5 pmol of each primer.

    RNA Extraction:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: Paragraph title: 2.2. RNA Extraction and Analysis ... The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    Amplification:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: To check for gDNA elimination, 1:10 dilutions of RT plus and minus reactions were used as a template for amplification of 7SL-RNA by qualitative PCR using DreamTaq (Life Technologies) and applying the manufacturer’s recommended conditions; negative amplification of RT minus reaction was used to verify the complete digestion of gDNA. .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    Nick Translation:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: PCR fragments were then 32P-labeled using the Nick Translation Kit (Hoffmann-La Roche, Basel, Switzerland). .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    Northern Blot:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: For Northern blot analysis, 6 μg of total RNA was electrophoresed and transferred onto Hybond-NX (GE Healtcare, Fairfield, CT, USA) membranes. .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    Quantitative RT-PCR:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ]. .. All PCR reactions were carried out in a final volume of 15 μL using 1 μL of diluted cDNA, 7.5 μL of 2X SYBR-Green (Bio-Rad) and 5 pmol of each primer.

    Electrophoresis:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: DNA extraction, manipulation and labelling, RNA electrophoresis, and blotting were carried out according to [ ]. .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    Polymerase Chain Reaction:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: PCR fragments were then 32P-labeled using the Nick Translation Kit (Hoffmann-La Roche, Basel, Switzerland). .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    Produced:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: Probes were produced by amplifying PCR gDNA fragments (0.3–0.5 kb in length) spanning the detected sequences. .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ].

    Real-time Polymerase Chain Reaction:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ]. .. All PCR reactions were carried out in a final volume of 15 μL using 1 μL of diluted cDNA, 7.5 μL of 2X SYBR-Green (Bio-Rad) and 5 pmol of each primer.

    Molecular Weight:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: .. The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ]. .. All PCR reactions were carried out in a final volume of 15 μL using 1 μL of diluted cDNA, 7.5 μL of 2X SYBR-Green (Bio-Rad) and 5 pmol of each primer.

    Software:

    Article Title: Unusual Novel SnoRNA-Like RNAs in Drosophila melanogaster
    Article Snippet: The size of RNAs was determined by using high range and low range RNA molecular weight markers (Life Technologies) Quantitative real-time RT-PCR (qRT-PCR) experiments were performed in triplicate using iQ5 Multicolor Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA) as previously described [ ]. .. All primer sequences were designed using Primer 3 software ( bioinfo.ut.ee/primer3-0.4.0/ ) [ ] and are available under request.

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    Thermo Fisher riboruler low range rna ladder
    Incubation of in vitro transcripts of CRISPR1, 2 and 3 with Cas6-1. ( A ) A total of 2.3 μM of CRISPR1 transcript is cleaved by 6.2 μM Cas6-1 by incubation for 1 h. ( B ) A total of 2 μM of CRISPR2 transcript is cleaved by 6.2 μM Cas6-1 by incubation for 1 h. ( C ) A total of 3.2 μM of CRISPR3 transcript is not cleaved by 6.5 μM Cas6-1 by incubation for 2 h. ( D ) The cleavage of 6.6 μM of the shorter CRISPR1* in vitro transcript by 5.8 μM Cas6-1 monitored over a time of 2 h 20 min. Since no finally mature crRNA ( 17 ) was detected, the in vivo presence of an unknown ribonuclease is suggested. All reactions were performed at 37°C in a reaction volume of 5 μl. <t>RNA</t> was separated by 8 M urea 10% PAGE and bands were visualized by ethidium bromide staining. Diamonds represent repeats and rectangles spacer sequences. Transcripts were not gel purified, explaining the appearance of fragments smaller than the full length transcripts of CRISPR1 and 2 in absence of Cas6-1. Eb1, elution buffer 1 used as negative control. Molecular markers: MN, Low Range ssRNA Ladder (NEB); MF, <t>RiboRuler</t> Low Range RNA Ladder (Thermo Fisher Scientific).
    Riboruler Low Range Rna Ladder, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Price from $9.99 to $1999.99
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    Incubation of in vitro transcripts of CRISPR1, 2 and 3 with Cas6-1. ( A ) A total of 2.3 μM of CRISPR1 transcript is cleaved by 6.2 μM Cas6-1 by incubation for 1 h. ( B ) A total of 2 μM of CRISPR2 transcript is cleaved by 6.2 μM Cas6-1 by incubation for 1 h. ( C ) A total of 3.2 μM of CRISPR3 transcript is not cleaved by 6.5 μM Cas6-1 by incubation for 2 h. ( D ) The cleavage of 6.6 μM of the shorter CRISPR1* in vitro transcript by 5.8 μM Cas6-1 monitored over a time of 2 h 20 min. Since no finally mature crRNA ( 17 ) was detected, the in vivo presence of an unknown ribonuclease is suggested. All reactions were performed at 37°C in a reaction volume of 5 μl. RNA was separated by 8 M urea 10% PAGE and bands were visualized by ethidium bromide staining. Diamonds represent repeats and rectangles spacer sequences. Transcripts were not gel purified, explaining the appearance of fragments smaller than the full length transcripts of CRISPR1 and 2 in absence of Cas6-1. Eb1, elution buffer 1 used as negative control. Molecular markers: MN, Low Range ssRNA Ladder (NEB); MF, RiboRuler Low Range RNA Ladder (Thermo Fisher Scientific).

    Journal: Nucleic Acids Research

    Article Title: Structural constraints and enzymatic promiscuity in the Cas6-dependent generation of crRNAs

    doi: 10.1093/nar/gkw786

    Figure Lengend Snippet: Incubation of in vitro transcripts of CRISPR1, 2 and 3 with Cas6-1. ( A ) A total of 2.3 μM of CRISPR1 transcript is cleaved by 6.2 μM Cas6-1 by incubation for 1 h. ( B ) A total of 2 μM of CRISPR2 transcript is cleaved by 6.2 μM Cas6-1 by incubation for 1 h. ( C ) A total of 3.2 μM of CRISPR3 transcript is not cleaved by 6.5 μM Cas6-1 by incubation for 2 h. ( D ) The cleavage of 6.6 μM of the shorter CRISPR1* in vitro transcript by 5.8 μM Cas6-1 monitored over a time of 2 h 20 min. Since no finally mature crRNA ( 17 ) was detected, the in vivo presence of an unknown ribonuclease is suggested. All reactions were performed at 37°C in a reaction volume of 5 μl. RNA was separated by 8 M urea 10% PAGE and bands were visualized by ethidium bromide staining. Diamonds represent repeats and rectangles spacer sequences. Transcripts were not gel purified, explaining the appearance of fragments smaller than the full length transcripts of CRISPR1 and 2 in absence of Cas6-1. Eb1, elution buffer 1 used as negative control. Molecular markers: MN, Low Range ssRNA Ladder (NEB); MF, RiboRuler Low Range RNA Ladder (Thermo Fisher Scientific).

    Article Snippet: Before loading onto denaturing 8 M urea 10% PAA gels, reactions were incubated at 95°C for 5 min. As size markers served the Low Range ssRNA Ladder from NEB (MN ), the RiboRuler Low Range RNA Ladder from Thermo Fisher Scientific (MF ) and the Low Molecular Weight Marker from Affymetrix (MA ).

    Techniques: Incubation, In Vitro, In Vivo, Polyacrylamide Gel Electrophoresis, Staining, Purification, Negative Control

    Purification and in vitro cleavage assay of Synechocystis WT Cas6-1 and mutated Cas6-1 proteins. (A) Coomassie-stained 15% SDS-PAGE of heterologously overexpressed and purified Cas6-1 protein variants. The expected molecular weight of Cas6-1_6xHis is ∼ 30 kDa. 66.67 pmol recombinant enzyme was loaded on each lane and PageRuler™ Prestained Protein Ladder (Thermo Fisher Scientific) served for protein size estimation. (B, C) In in vitro cleavage assays both the (B) CRISPR1 RNA oligo repeat sequence and the (C) pre-acrRNA cassette is processed with variable efficiency by the purified Cas6-1 protein variants shown in (A). After incubation at 30°C the RNA fragments were electrophoretically size separated on a 15% PAA sequencing gel and visualized with SYBR® Gold Nucleic Acid Gel Stain. The processing results in a pattern of RNA fragments of distinct lengths. Cleavage assays were conducted with a three-fold excess (B) or a 1:1 ratio (C) of protein to RNA-substrate in cleavage buffer and incubated at 30°C for the indicated time. Expected RNA fragments and their corresponding sizes are illustrated by cartoons. Another Cas6-1 WT protein sample, which was overexpressed and purified independently, is indicated by an asterisk. Negative controls were complemented with 1 μL of H 2 O or 1 μL eluate of a mock control. Two additional nucleotides resulting from in vitro transcription were added to the 215 nt long acrRNA yielding a 217 nt long pre-acrRNA. For RNA size estimation a CRISPR1 repeat oligo ladder (C1L), RiboRuler low range RNA ladder (Thermo Fisher Scientific), low range ssRNA ladder or microRNA marker (both from New England Biolabs) were used.

    Journal: RNA Biology

    Article Title: Biochemical analysis of the Cas6-1 RNA endonuclease associated with the subtype I-D CRISPR-Cas system in Synechocystis sp. PCC 6803

    doi: 10.1080/15476286.2018.1447742

    Figure Lengend Snippet: Purification and in vitro cleavage assay of Synechocystis WT Cas6-1 and mutated Cas6-1 proteins. (A) Coomassie-stained 15% SDS-PAGE of heterologously overexpressed and purified Cas6-1 protein variants. The expected molecular weight of Cas6-1_6xHis is ∼ 30 kDa. 66.67 pmol recombinant enzyme was loaded on each lane and PageRuler™ Prestained Protein Ladder (Thermo Fisher Scientific) served for protein size estimation. (B, C) In in vitro cleavage assays both the (B) CRISPR1 RNA oligo repeat sequence and the (C) pre-acrRNA cassette is processed with variable efficiency by the purified Cas6-1 protein variants shown in (A). After incubation at 30°C the RNA fragments were electrophoretically size separated on a 15% PAA sequencing gel and visualized with SYBR® Gold Nucleic Acid Gel Stain. The processing results in a pattern of RNA fragments of distinct lengths. Cleavage assays were conducted with a three-fold excess (B) or a 1:1 ratio (C) of protein to RNA-substrate in cleavage buffer and incubated at 30°C for the indicated time. Expected RNA fragments and their corresponding sizes are illustrated by cartoons. Another Cas6-1 WT protein sample, which was overexpressed and purified independently, is indicated by an asterisk. Negative controls were complemented with 1 μL of H 2 O or 1 μL eluate of a mock control. Two additional nucleotides resulting from in vitro transcription were added to the 215 nt long acrRNA yielding a 217 nt long pre-acrRNA. For RNA size estimation a CRISPR1 repeat oligo ladder (C1L), RiboRuler low range RNA ladder (Thermo Fisher Scientific), low range ssRNA ladder or microRNA marker (both from New England Biolabs) were used.

    Article Snippet: The low range ssRNA ladder (ssRNA marker) (New England Biolabs), the RiboRuler low range RNA ladder (Thermo Fisher Scientific) and the microRNA marker (New England Biolabs) served as RNA ladders for size estimation.

    Techniques: Purification, In Vitro, Cleavage Assay, Staining, SDS Page, Molecular Weight, Recombinant, Sequencing, Incubation, Marker

    In vivo cleavage assay of Cas6-1 protein variants and pre-acrRNA. Northern Blot analysis using a radioactively labeled transcript probe spanning the entire pre-acrRNA cassette. The RiboRuler low range RNA ladder was used as a size marker. Cas6-1 expression was controlled by the copper inducible petE , ], whereas acrRNA was zinc inducible by the ziaA ]. The smallest detectable processing intermediate of 73 nt in size is marked by asterisks. WT Cas6-1 and the R67A protein variant show comparable cleavage performance indicating that R67A is not involved in the active site or RNA binding of the enzyme. In contrast, the remaining mutated proteins show distinct acrRNA accumulation and lack acrRNA derived cleavage intermediates.

    Journal: RNA Biology

    Article Title: Biochemical analysis of the Cas6-1 RNA endonuclease associated with the subtype I-D CRISPR-Cas system in Synechocystis sp. PCC 6803

    doi: 10.1080/15476286.2018.1447742

    Figure Lengend Snippet: In vivo cleavage assay of Cas6-1 protein variants and pre-acrRNA. Northern Blot analysis using a radioactively labeled transcript probe spanning the entire pre-acrRNA cassette. The RiboRuler low range RNA ladder was used as a size marker. Cas6-1 expression was controlled by the copper inducible petE , ], whereas acrRNA was zinc inducible by the ziaA ]. The smallest detectable processing intermediate of 73 nt in size is marked by asterisks. WT Cas6-1 and the R67A protein variant show comparable cleavage performance indicating that R67A is not involved in the active site or RNA binding of the enzyme. In contrast, the remaining mutated proteins show distinct acrRNA accumulation and lack acrRNA derived cleavage intermediates.

    Article Snippet: The low range ssRNA ladder (ssRNA marker) (New England Biolabs), the RiboRuler low range RNA ladder (Thermo Fisher Scientific) and the microRNA marker (New England Biolabs) served as RNA ladders for size estimation.

    Techniques: In Vivo, Cleavage Assay, Northern Blot, Labeling, Marker, Expressing, Variant Assay, RNA Binding Assay, Derivative Assay