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Related Concept Videos

pre-mRNA Processing02:01

pre-mRNA Processing

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In eukaryotic cells, transcripts made by RNA polymerase are modified and processed before exiting the nucleus. Unprocessed RNA is called precursor mRNA or pre-mRNA to distinguish it from mature mRNA.
Once about 20-40 ribonucleotides have been joined together by RNA polymerase, a group of enzymes adds a “cap” to the 5’ end of the growing transcript. In this process, a 5’ phosphate is replaced by modified guanosine that has a methyl group attached to it (7-Methyl...
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Regulated mRNA Transport02:22

Regulated mRNA Transport

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In eukaryotes, transcription and translation are compartmentalized; an mRNA is first synthesized in the nucleus and then selectively transported to the cytoplasm for protein synthesis. Before transport, a pre-mRNA undergoes several steps of post-transcriptional modifications including splicing, 5' capping, and the addition of a poly-adenine tail. Various proteins bind to the pre-mRNA during these modifications. The mRNA transport takes place with the help of multiple proteins playing...
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Nuclear Export of mRNA02:31

Nuclear Export of mRNA

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Before mRNAs are exported to the cytoplasm, it is crucial to check each mRNA for structural and functional integrity. Eukaryotic cells use several different mechanisms, collectively known as mRNA surveillance, to look for irregularities in mRNAs. Irregular or aberrant mRNA are rapidly degraded by various enzymes. If a defective mRNA escapes the surveillance, it would be translated into a protein which would either be non-functional or not function properly. One of the primary irregularities in...
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mRNA Stability and Gene Expression02:51

mRNA Stability and Gene Expression

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The structure and stability of mRNA molecules regulates gene expression, as mRNAs are a key step in the pathway from gene to protein. In eukaryotes, the half-life of mRNA varies from a few minutes up to several days. mRNA stability is essential in growth and development. The absence of the proteins regulating its stability, such as tristetraprolin in mice, can cause systemic issues, including bone marrow overgrowth, inflammation, and autoimmunity.
Cis-acting Elements involved in mRNA stability
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Nonsense-mediated mRNA Decay02:27

Nonsense-mediated mRNA Decay

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The Upf proteins that carry out nonsense-mediated decay (NMD) are found in all eukaryotic organisms, including humans. Each protein has an individual role, but they need to work in collaboration. Upf1 is an ATP-dependent RNA helicase that unwinds the RNA helix. Because Upf1 can unwind any RNA, Upf2 and Upf3 are required to help Upf1 discriminate between nonsense and normal mRNAs.
Usually, Upf3 binds to an Exon Junction Complex (EJC) at mRNA splice sites. If a ribosome fully translates the mRNA,...
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Power Distribution in Three-phase and Single Phase Circuits01:17

Power Distribution in Three-phase and Single Phase Circuits

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Power distribution within electrical circuits is a foundational aspect of residential and industrial energy systems. While single-phase power is common in residential settings, three-phase power is the standard for industrial environments with heavy machinery. Each system is different and has advantages, and it's crucial to understand the underlying principles of power distribution and material efficiency.
Single-Phase Power Distribution:
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Single Read and Paired End mRNA-Seq Illumina Libraries from 10 Nanograms Total RNA
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Single-Cell mRNA-Seq Using the Fluidigm C1 System and Integrated Fluidics Circuits.

Haibiao Gong1, Devin Do1, Ramesh Ramakrishnan2,3

  • 1Fluidigm Corporation, South San Francisco, CA, USA.

Methods in Molecular Biology (Clifton, N.J.)
|May 17, 2018
PubMed
Summary

This study details a single-cell messenger RNA sequencing (mRNA-seq) workflow using microfluidics for high-throughput cell analysis. The method provides valuable full-length mRNA data for understanding gene regulatory networks.

Keywords:
Gene expressionIntegrated fluidic circuitSingle-cellmRNA-seq

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Area of Science:

  • Molecular Biology
  • Genomics
  • Biotechnology

Background:

  • Single-cell mRNA sequencing (mRNA-seq) is crucial for dissecting gene expression and regulatory networks.
  • Microfluidics offers advantages for single-cell analysis due to small reaction volumes and automation capabilities.

Purpose of the Study:

  • To describe a comprehensive workflow for single-cell mRNA-seq.
  • To demonstrate the utility of the C1 Integrated Fluidic Circuit (IFC) for isolating and processing up to 96 single cells.
  • To highlight the generation of full-length mRNA information.

Main Methods:

  • Utilized the C1 IFC for automated single-cell isolation and processing.
  • Performed on-chip lysis, reverse transcription, and preamplification PCR.
  • Executed off-chip sequencing library preparation.

Main Results:

  • Successfully implemented a workflow for single-cell mRNA-seq using C1 IFC.
  • Generated full-length mRNA sequence data from up to 96 individual cells.
  • Demonstrated the value of full-length mRNA information over 3' end counting methods.

Conclusions:

  • The described workflow enables efficient and high-throughput single-cell mRNA-seq.
  • The C1 IFC platform facilitates comprehensive gene expression analysis at the single-cell level.
  • Full-length mRNA sequencing provides richer data for biological insights compared to 3' end methods.