Molecular Diagnostics Chapter 6 EXAM GRADED A+

Molecular Diagnostics Chapter 6 EXAM GRADED A+ PCR - ANSWER(polymerase chain reaction) a method used to rapidly make multiple copies of a specific segment of DNA; can be used to make millions of copies of DNA from a very small amount of DNA Target amplification - ANSWERTarget: Specific sequences or regions that are detected in an analytical procedure. Amplification: Replication/copying Target amplification: replication of a specific target region of DNA. dNTPs - ANSWERdeoxynucleotide triphosphates What are the three stages of PCR? What are the temperatures and time of each stage? - ANSWERDenature-90-96 degrees celsius, 20-60s Anneal-50-70 degrees celsius, 20-90s Extension-68-75 degrees, 10-60s What are the typical components of a PCR reaction? - ANSWER-primer to prime target for replication -dNTP's for extension of new strand -KCl-salt helps DNA hybridize -Tris buffer to maintain pH -MgCl2- cation requirement for polymerase -polymerase -template/target The forward primer binds on which DNA template orientation 5'->3' or 3'->5'? - ANSWER3'-5' as the forward primer is oriented in the 5' to 3' direction The reverse primer binds on which DNA template orientation 5'->3' or 3'->5'? - ANSWER5' to 3' Which step of PCR determines the specificity of the reaction? - ANSWERprimer annealing to template strand because determines what will be duplicated primer design - ANSWER• Primers should be 20 - 30 base pairs long (too short = not very specific, too long = difficult to bind to D