DNA Structure and Nucleases Summary

  • Meselson and Stahl Paper: Due at the next class along with Quiz #5.

  • DNA Structure:

    • Stores genetic information in its base sequence.
    • Easily copied, crucial for replication.

  • Nucleases:

    • Enzymes that degrade nucleic acids to protect against foreign DNA.
    • Two types:
    1. Exonucleases: Digest DNA from the ends.
    2. Endonucleases: Cut DNA in the middle, also known as restriction endonucleases for specific sequences.

  • Restriction Endonucleases Examples:

    • Eco RI (from Escherichia coli): Target sequence - GAA TTC.
    • Bam HI (from Bacillus amyloliquefaciens): Target sequence - GGATCC.
    • Hind III (from Haemophilus influenzae): Target sequence - AAGCTT.
    • All listed sequences are palindromic (same forwards and backwards).

  • Plasmids:

    • Small circular DNA molecules separate from chromosomal DNA.
    • Can confer antibiotic resistance and be exchanged between bacteria.

  • DNA Fragmentation:

    • One target site = 1 fragment.
    • Two target sites = 2 fragments (e.g., Eco RI on a 3kb plasmid results in two fragments of 2kb and 1kb).

  • Separation of DNA Fragments:

    • Through electrophoresis using agarose gels; smaller fragments migrate faster due to charge and size.

  • RFLP Analysis:

    • Uses restriction fragment length polymorphism for identification.
    • Unique patterns help in forensic applications (e.g., O.J. Simpson case).

  • DNA Hybridization:

    • Measures similarity, where DNA strands can bind (hybridize) if they are complimentary.
    • Hybridization is influenced by temperature and number of complementary bases.