Primer extensionPrimer extension• * This labelling technique uses random

oligonucleotides (usually hexadeoxyribonucleotide molecules- sequences of six deoxynucleotides) to primer synthesis of a DNA strand by DNA polymerase.

• * The DNA to be labelled is denaturated by heating, and the oligonucleotide primer annealed to the single stranded DNAs.

•The klenowThe klenow fragment of DNA fragment of DNA polymerase can synthesize a polymerase can synthesize a copy of the template, primed copy of the template, primed from the 3-hydroxyl group of the from the 3-hydroxyl group of the oligonucleotide.oligonucleotide.

* If a labelled dNTP is * If a labelled dNTP is incorported, DNA of very high incorported, DNA of very high specific activity is produced. specific activity is produced. (Fig (Fig 12)12)

Nucleic acid hybridizationNucleic acid hybridization

•The complementary feature of DNA base-pairing The complementary feature of DNA base-pairing can be used to provide information a bout the can be used to provide information a bout the sequence complexity of the DNA.sequence complexity of the DNA.

•If a DNA duplex is denaturated by heating until If a DNA duplex is denaturated by heating until the strands separate, the complementary strands the strands separate, the complementary strands will be renaturated on cooling . will be renaturated on cooling . (Fig 13)(Fig 13)