Difference between revisions of "Part:BBa K4361201"
Line 3: Line 3:
 
<partinfo>BBa_K4361201 short</partinfo>
 
<partinfo>BBa_K4361201 short</partinfo>
  
−
For the site-directed mutagenesis (SDM) of our protein of interest, BlcR, multiple sets of back-to-back primers were designed. The 5' ends of these primers anneal to adjacent nucleotides such that the full plasmid containing the protein ([[Part:BBa_K4361106]]) can be replicated. Each of these sets of primers contains a single reverse primer, denoted as "primer R<i>n</i>" with <i>n</i> being the number of the set, in this case primer R1 ([[Part:BBa_K4361200]]), and a number of forward primers including this BioBrick. This set covers amino acids 36 to 39, although 36 is not represented in our BioBricks due to that SDM experiment being unsuccessful.<br>
+
For the site-directed mutagenesis (SDM) of our protein of interest, BlcR, multiple sets of back-to-back (BTB) primers were designed. The 5' ends of these primers anneal to adjacent nucleotides such that the full plasmid containing the protein ([[Part:BBa_K4361106]]) can be replicated. Each of these numbered sets of primers contains a single reverse primer (denoted as R<i>n</i> with <n> being the number of the set) and a varying number of forward primers (denoted as F<i>n</i>.<i>m</i>). Every set covers a small range of amino acids in the DNA-binding domain of BlcR, with each position being mutated to different residues by different forward-facing primers. Do note, however, that while a large number of primers have been designed, sequencing showed that the mutagenesis protocol was not successful for all variants. Therefore, only the primers that resulted in the desired mutants will be described in our collection of BioBricks.<br>
 
<br>
 
<br>
 +
The primer sets are divided as follows:
 +
<ul>
 +
  <li>Set 1, residues 36 to 39
 +
    <ul>
 +
      <li>[[Part:BBa_K4361200]]:
 +
 
This primer contains a mutation that changes the codon for aspartic acid (D, GAC) in position 37 to that of arginine (R, CGC). <br>
 
This primer contains a mutation that changes the codon for aspartic acid (D, GAC) in position 37 to that of arginine (R, CGC). <br>
 
<br>
 
<br>

Revision as of 12:36, 14 September 2022


BlcR BTB primer F1.1 D37R

For the site-directed mutagenesis (SDM) of our protein of interest, BlcR, multiple sets of back-to-back (BTB) primers were designed. The 5' ends of these primers anneal to adjacent nucleotides such that the full plasmid containing the protein (Part:BBa_K4361106) can be replicated. Each of these numbered sets of primers contains a single reverse primer (denoted as Rn with <n> being the number of the set) and a varying number of forward primers (denoted as Fn.m). Every set covers a small range of amino acids in the DNA-binding domain of BlcR, with each position being mutated to different residues by different forward-facing primers. Do note, however, that while a large number of primers have been designed, sequencing showed that the mutagenesis protocol was not successful for all variants. Therefore, only the primers that resulted in the desired mutants will be described in our collection of BioBricks.

The primer sets are divided as follows:

  • Set 1, residues 36 to 39
    • Part:BBa_K4361200: This primer contains a mutation that changes the codon for aspartic acid (D, GAC) in position 37 to that of arginine (R, CGC).

      Sequence and Features

      Assembly Compatibility:
      • 10
        COMPATIBLE WITH RFC[10]
      • 12
        COMPATIBLE WITH RFC[12]
      • 21
        COMPATIBLE WITH RFC[21]
      • 23
        COMPATIBLE WITH RFC[23]
      • 25
        COMPATIBLE WITH RFC[25]
      • 1000
        COMPATIBLE WITH RFC[1000]