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Related Concept Videos

PCR01:32

PCR

Overview
RACE - Rapid Amplification of cDNA Ends02:35

RACE - Rapid Amplification of cDNA Ends

Rapid Amplification of cDNA Ends, or RACE, is one of the most effective methods to obtain a full-length cDNA from an mRNA sequence between a known internal region to the unknown sequence at the 5’ or 3’ end. The unknown region is cloned in the cDNA by a gene-specific primer that binds the known end, and a hybrid primer that attaches a predefined anchor sequence to the unknown end of the cDNA. The sequence in between is amplified by PCR with an anchor primer and a gene-specific primer.
Since the...
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Genome-wide Association Studies-GWAS

Genome-wide association studies or GWAS are used to identify whether common SNPs are associated with certain diseases. Suppose specific SNPs are more frequently observed in individuals with a particular disease than those without the disease. In that case, those SNPs are said to be associated with the disease. Chi-square analysis is performed to check the probability of the allele likely to be associated with the disease.
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Sanger Sequencing

DNA sequencing is a fundamental technique that is routinely used in the biological sciences. This method can be applied to a range of questions at different scales - from the sequencing of a cloned DNA fragment or the study of a mutation in a gene up to whole-genome sequencing. However, despite the widespread use of sequencing today, it was not until 1977 that Fredrick Sanger and his collaborators developed the chain-termination method to decode DNA sequences. It relies on the separation of a...
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.
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Related Experiment Video

Updated: Jul 5, 2026

Amplification of Near Full-length HIV-1 Proviruses for Next-Generation Sequencing
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Amplification of sequences from affected individuals.

H C Watkins1

  • 1Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts, USA.

Current Protocols in Human Genetics
|April 23, 2008
PubMed
Summary

This study details methods for DNA sequencing in genetic disorder patients, using polymerase chain reaction (PCR) to amplify low-copy number targets from patient samples. These protocols enable efficient gene analysis from accessible lymphocytes or genomic DNA.

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Last Updated: Jul 5, 2026

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Wild-type Blocking PCR Combined with Direct Sequencing as a Highly Sensitive Method for Detection of Low-Frequency Somatic Mutations
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Wild-type Blocking PCR Combined with Direct Sequencing as a Highly Sensitive Method for Detection of Low-Frequency Somatic Mutations

Published on: March 29, 2017

Area of Science:

  • Molecular Biology
  • Genetics
  • Biotechnology

Background:

  • Genetic disorders require accurate DNA sequencing for diagnosis and research.
  • Analyzing DNA from patient samples, especially those with low target sequence copy numbers, presents technical challenges.

Purpose of the Study:

  • To describe robust methods for obtaining DNA sequences from individuals with genetic disorders.
  • To provide protocols for amplifying and analyzing target DNA sequences, including those present at low concentrations.

Main Methods:

  • Utilizing polymerase chain reaction (PCR) for amplification of target DNA sequences.
  • Harvesting mRNA from peripheral lymphocytes for gene expression analysis (Basic Protocol 1).
  • Modifying PCR conditions for enhanced mutation analysis and sequencing, and direct genomic DNA analysis (Basic Protocol 2).

Main Results:

  • Successful amplification of low-copy number target sequences from patient samples.
  • Efficient screening of coding sequences using mRNA from peripheral lymphocytes.
  • Facilitation of mutation analysis and sequencing through optimized PCR conditions.

Conclusions:

  • The described methods provide effective strategies for DNA sequencing in genetic disorder research.
  • These protocols are adaptable for analyzing various gene regions, including exons and noncoding sequences.
  • The techniques offer efficient sample utilization and enhanced analysis capabilities for genetic studies.