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Related Experiment Videos

A 3.5 genome equivalent multi access YAC library: construction, characterisation, screening and storage.

R Anand1, J H Riley, R Butler

  • 1ICI Diagnostics, Northwich, Cheshire, UK.

Nucleic Acids Research
|April 25, 1990
PubMed
Summary

A new yeast artificial chromosome (YAC) library was created from human DNA, offering extensive genome coverage. This resource facilitates the isolation of specific DNA fragments for research purposes.

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Area of Science:

  • Genomics
  • Molecular Biology
  • Biotechnology

Background:

  • Human lymphoblastoid cell lines are valuable for genetic studies.
  • Yeast artificial chromosomes (YACs) are essential tools for constructing large genomic libraries.
  • Efficient storage and accessibility of genomic libraries are crucial for multi-user research.

Purpose of the Study:

  • To construct a comprehensive yeast artificial chromosome (YAC) primary gridded library.
  • To ensure high genome coverage and long-term stability of the library.
  • To validate the library's utility for isolating specific human DNA sequences.

Main Methods:

  • Construction of a gridded YAC library using human lymphoblastoid (48,XXXX) cell line DNA.
  • Characterization of average YAC size and genome coverage.

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  • Storage of the library at -70°C as gridded filters and glycerol suspensions.
  • Screening the library using colony hybridization and PCR with single-copy probes.

    • Main Results:

    • A primary gridded YAC library of 35,000 clones was successfully constructed.
    • The average YAC size was approximately 350kb, providing >3.5x genome coverage.
    • Screening with 14 single-copy probes yielded 1-5 YAC clones per probe.

    Conclusions:

    • The developed YAC library is a robust and valuable resource for human genomics research.
    • The library's construction and storage methods ensure prolonged multi-user potential.
    • The successful isolation of YAC clones demonstrates the library's effectiveness for gene mapping and analysis.