In Polymerase Chain Reaction (PCR) primers are used to provide:
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Remember: D elongation always proceeds from $5'$ to $3'$.
A free $3'$-OH group is indispensable for phosphodiester bond formation during D replication and PCR amplifications.
Step 1: Understanding the Question:
The question asks about the fundamental role of primers in the Polymerase Chain Reaction (PCR), specifically identifying which chemical group at the end of the primer allows D polymerase to attach new nucleotides. Step 2: Key Formula or Approach:
All D polymerases synthesize new D strands exclusively in the $5' \to 3'$ direction.
The catalytic core of D polymerase requires a free hydroxyl group ($-OH$) at the $3'$ position of the deoxyribose sugar to perform a nucleophilic attack on the $\alpha$-phosphate of an incoming deoxyribonucleoside triphosphate (dNTP). Step 3: Detailed Explation:
• Role of Primers in PCR: Primers are short, synthetic single-stranded oligonucleotides (usually 18 to 30 nucleotides long) designed to complement specific target sequences on template D.
• Requirement for Polymerization: D polymerases cannot initiate D synthesis $de\ novo$; they can only add nucleotides to an existing pre-formed polynucleotide strand.
• Chemical Interaction: The $3'$-end of the hybridized primer presents a free $3'$-OH group. During strand extension, the oxygen atom of this $3'$-OH group carries out a nucleophilic attack on the $\alpha$-phosphorus atom of the incoming dNTP, forming a phosphodiester bond and releasing pyrophosphate ($PP_i$).
• 5' and Phosphate Groups: The $5'$ end of synthetic primers typically possesses a $5'$-hydroxyl or $5'$-phosphate group, but this end is not involved in chain elongation. Therefore, $3'$-PO$_4^-$ or $5'$-PO$_4^-$ cannot serve as starting points for D extension. Step 4: Fil Answer:
Synthetic primers in PCR provide the necessary free $3'$-OH group required by D polymerase to extend the target D sequence.
