Difference between revisions of "Part:BBa K3582500"
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pET28a(+) is a novel expression plasmid generally used in E-coli strains for protein production purposes. It has a kanamycin resistance gene as a selection marker. Plasmid sequence consists of the T7 promoter and terminator sequences as well as lac operon sequence and lac promoter for protein expression induction. Apart from this, it also has well-defined operator, promoter, MCS and tag sequences precisely inbuilt. | pET28a(+) is a novel expression plasmid generally used in E-coli strains for protein production purposes. It has a kanamycin resistance gene as a selection marker. Plasmid sequence consists of the T7 promoter and terminator sequences as well as lac operon sequence and lac promoter for protein expression induction. Apart from this, it also has well-defined operator, promoter, MCS and tag sequences precisely inbuilt. | ||
− | The normal pet28a(+) vector contains several unnecessary components like 6Xhis tag, thrombin site etc. that could interfere with protein expression and purification. Identifying a proper insertion site becomes difficult. The normal version does not even satisfy the | + | The normal pet28a(+) vector contains several unnecessary components like 6Xhis tag, thrombin site etc. that could interfere with protein expression and purification. Identifying a proper insertion site becomes difficult. The normal version does not even satisfy the RFC10 criteria. |
− | This version of Pet28a(+) vector is stringently modified by removing all the unnecessary tags and non required regions. The insertion site is aptly annotated with a start and a stop codon where the insertion of the ORF can be made. Most importantly | + | This version of Pet28a(+) vector is stringently modified by removing all the unnecessary tags and non required regions. The insertion site is aptly annotated with a start and a stop codon where the insertion of the ORF can be made. Most importantly RFC10 criteria are satisfied and all of the other features of the vector are conserved and undisturbed. |
Revision as of 17:29, 24 October 2020
Modified Pet28a(+) expression vector
pET28a(+) is a novel expression plasmid generally used in E-coli strains for protein production purposes. It has a kanamycin resistance gene as a selection marker. Plasmid sequence consists of the T7 promoter and terminator sequences as well as lac operon sequence and lac promoter for protein expression induction. Apart from this, it also has well-defined operator, promoter, MCS and tag sequences precisely inbuilt.
The normal pet28a(+) vector contains several unnecessary components like 6Xhis tag, thrombin site etc. that could interfere with protein expression and purification. Identifying a proper insertion site becomes difficult. The normal version does not even satisfy the RFC10 criteria.
This version of Pet28a(+) vector is stringently modified by removing all the unnecessary tags and non required regions. The insertion site is aptly annotated with a start and a stop codon where the insertion of the ORF can be made. Most importantly RFC10 criteria are satisfied and all of the other features of the vector are conserved and undisturbed.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 105
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 137
Illegal NgoMIV site found at 1725
Illegal NgoMIV site found at 1885
Illegal NgoMIV site found at 4932 - 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI site found at 2805