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Related Concept Videos

RNA-seq03:21

RNA-seq

RNA sequencing, or RNA-Seq, is a high-throughput sequencing technology used to study the transcriptome of a cell. Transcriptomics helps to interpret the functional elements of a genome and identify the molecular constituents of an organism. Additionally, it also helps in understanding the development of an organism and the occurrence of diseases. 
Before the discovery of RNA-seq, microarray-based methods and Sanger sequencing were used for transcriptome analysis. However, while microarray-based...
Next-generation Sequencing03:00

Next-generation Sequencing

The first human genome sequencing project cost $2.7 billion and was declared complete in 2003, after 15 years of international cooperation and collaboration between several research teams and funding agencies. Today, with the advent of next-generation sequencing technologies, the cost and time of sequencing a human genome have dropped over 100 fold.
Next-Generation Sequencing Methods
Although all next-generation methods use different technologies, they all share a set of standard features.

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Multiplexed Single Cell mRNA Sequencing Analysis of Mouse Embryonic Cells
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Generating barcoded libraries for multiplex high-throughput sequencing.

Michael Knapp1, Mathias Stiller, Matthias Meyer

  • 1Allan Wilson Centre for Molecular Ecology and Evolution, Department of Anatomy and Structural Biology, University of Otago, Dunedin 9016, New Zealand. michael.knapp@otago.ac.nz

Methods in Molecular Biology (Clifton, N.J.)
|January 13, 2012
PubMed
Summary
This summary is machine-generated.

Molecular barcoding enables optimal use of high-throughput sequencing for multiple samples. This study presents two novel polymerase chain reaction (PCR) methods for creating barcoded libraries from low-quantity ancient DNA.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Bioinformatics

Background:

  • High-throughput sequencing (HTS) platforms offer powerful analytical capabilities.
  • Molecular barcoding is crucial for multiplexing samples in HTS studies.
  • Ancient DNA (aDNA) studies often yield low DNA concentrations, posing challenges for library preparation.

Purpose of the Study:

  • To develop and optimize molecular barcoding strategies for low-input DNA samples.
  • To adapt existing barcoding techniques for the specific requirements of ancient DNA research.
  • To enhance the efficiency of generating barcoded sequencing libraries from challenging biological materials.

Main Methods:

  • Two distinct molecular barcoding approaches were developed.
  • Methods involved the incorporation of short recognition sequences (barcodes) into sequencing libraries.
  • Techniques were optimized for low copy number DNA extracts and amplification products.

Main Results:

  • Successful generation of barcoded sequencing libraries from low-input DNA.
  • Demonstrated applicability of the methods to ancient DNA extracts.
  • Provided optimized protocols for PCR-based barcoding.

Conclusions:

  • The presented molecular barcoding approaches are effective for low-input DNA samples.
  • These methods facilitate the use of HTS for ancient DNA studies.
  • Optimized barcoding strategies improve data yield and quality in challenging genomic research.