18. Biotechnology

Introduction to Polymerase Chain Reaction

Different component of PCR master mix

 determines how many cycles of the reaction are needed to obtain a sufficient amount of amplified DNA 

 No. Primers provide specificity to the reaction. 

 determines the number of tandem repeats in a genome 

 determines which segment of the genome will be amplified 

Consider the role of primers in DNA replication.

In the polymerase chain reaction (PCR), the sequence of bases in the primers is important because it __________.

 No. Plants are not a source of reverse transcriptase. 

 No. Fungi are not a source of reverse transcriptase. 

 Cultured phage-infected mammalian cells 

 No. Such cells are not a source of reverse transcriptase. 

 Either retroviruses or cultured phage-infected mammalian cells 

How does the HIV virus replicate in a host cell?

What is the source of the reverse transcriptase used in recombinant DNA technology?

Amplify a single gene or small sequence of DNA.

 Insert foreign DNA into a host cell where it can be replicated.

Cut segments of DNA creating complimentary sticky ends.

PCR (Polymerase Chain Reaction)

Components of a Polymerase Chain Reaction

PCR Ingredients

Doubles the template sequence of DNA with each cycle.

Makes a single copy of the template sequence of DNA with each cycle.

Is used to determine the sequence of a region of DNA.

 alter the bacteria so that they can splice RNA 

 No. This would involve engineering the bacteria to differentiate among the introns and exons, to cut out the introns, and to splice together the exons. There is a much easier way. 

 use a nucleic acid probe to find a gene without introns 

 No. Most eukaryotic genes have introns. 

 use a phage to insert the desired gene into a bacterium 

 No. This use of a phage will not eliminate the introns. 

 use a restriction enzyme to remove introns from the gene 

 No. Restriction enzymes cut at specific recognition sites that are unlikely to be in the right places for removing all the introns from a particular gene. 

 work backward from mRNA to make a version of the gene without introns 

When are introns normally removed?

Because eukaryotic genes contain introns, their transcripts cannot be translated by bacteria, which lack RNA-splicing machinery. But if you want to engineer a bacterium to produce a eukaryotic protein, you can synthesize a gene without introns. A good way to do this is to __________.

 No. DNA ligase creates sugar-phosphate bonds that connect fragments of DNA, but it is not used in PCR. 

 No. Reverse transcriptase catalyzes the formation of cDNA from an RNA template; PCR makes DNA copies of a DNA segment. 

 No. Restriction enzymes cut DNA at specific recognition sites and are not used in PCR. 

 No. Although DNA polymerase is the type of enzyme used to amplify DNA in PCR, human DNA polymerase would be denatured by the high temperatures used at the start of each PCR cycle. 

PCR can quickly amplify any specific DNA segment within one or many DNA molecules in a test tube. What enzyme serves to "amplify" segments of existing DNA?

Which of the following enzymes is key to the automation of polymerase chain reactions (PCR)?

Polymerase Chain Reaction (PCR) Protocol

PCR - Polymerase Chain Reaction Simplified

Why do we use PCR?

What is Polymerase Chain Reaction? | PCR Explained

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