BIOLOGY 172 Lecture Notes - Lecture 21: Integrase, Stop Codon, Sanger Sequencing
Document Summary
Used to determine the nucleotide sequence of an unknown stretch of. Most widely used method = sanger sequencing: relies on dideoxynucleotides (ddntps, necessary for sequencing: Dntps used by dna polymerase to make dna. Has a h(cid:455)d(cid:396)o(cid:454)(cid:455)l g(cid:396)oup o(cid:374) the 3" (cid:272)a(cid:396)(cid:271)o(cid:374), allo(cid:449)i(cid:374)g this end to be elongated. La(cid:272)ks the 3" h(cid:455)d(cid:396)o(cid:454)(cid:455)l g(cid:396)oup p(cid:396)ese(cid:374)t i(cid:374) dntps; cannot be regulated by dna polymerase: the process: Within a test tube are many molecules of template, primer, nucleotides, and polymerase. Dna pol(cid:455)(cid:373)e(cid:396)ase adds (cid:272)o(cid:373)ple(cid:373)e(cid:374)ta(cid:396)(cid:455) (cid:271)ases to the 3" e(cid:374)d. Fo(cid:396) ea(cid:272)h lette(cid:396) (cid:271)ase, the(cid:396)e"s a dntp a(cid:374)d a ddntp version chance determine which one binds. Each ddntp is labelled with a different fluorescent dye for identification purposes. If a dntp is added, dna polymerase continues extending the strand. If a ddntp is added, chain is terminated because dna polymerase cannot add bases to it: end result: very many dna chains varying in size and color of fluorescent marker present at its end.
