Gene Isolation and Manipulation 4. Ligase is an essential enzyme within all cells that seals breaks in the phosphate-sugar backbone of DNA. During DNA replication it joins Okazaki fragments to create a continuous strand, and in cloning, it is used to join the various DNA fragments with the vector. If it was not added, the vector and cloned DNA would simply fall apart. 6. Each cycle takes five minutes and doubles the DNA. In one hour there would be 12 cycles so the DNA would be amplified 2 12 = 4096-fold amplification. 8. plasmid (15–20 kb) < cosmid (35–45 kb) < BAC (150–300 kb) < YAC (300 kb and larger) 15. The commercial cloning of insulin was into bacteria. Bacteria are not capable of processing introns. Genomic DNA would include the introns, while cDNA is a copy of processed (and thus intron-free) mRNA. 16. This problem assumes a random and equal distribution of nucleotides. Alu I ( 1 / 4 ) 4 = on average, once in every 256 nucleotide pairs Eco RI ( 1 / 4 ) 6 = on average, once in every 4096 nucleotide pairs
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This note was uploaded on 09/25/2010 for the course BIOL 3301 taught by Professor Gunaratne during the Fall '05 term at University of Houston.