Part:BBa_K1898450
Strong promoter + Strong RBS + CRYAB + 10x Histidine tag + Double terminator
BBa_K880005 is used to provide a strong promoter and strong rbs to maximize protein output. The ORF includes cryab, a crystallin protein exists in the lens and other tissues, and 10x Histidine tag for protein purification. BBa_B0015 consists of a double terminator to end transcription.
BBa_K880005 consists of a strong promoter and strong RBS and was used to maximize protein production. This construct codes for CRYAB (crystallin alpha B) and a 10x Histidine-tag. CRYAB is a crystallin protein abundant in the lens and 10x Histidine-tag is used for protein purification. BBa__B0015 is a double terminator used to stop transcription.
Usage and Biology
This composite part can be used for teams who want to purify crystallin protein. Recent studies have been done on alpha B crystallin and neurological diseases such as Alzhiemer and Parkinson's disease. [1]
For our project, this construct allows us to 1) test our protein purification method and 2) see the effect of oxidative stress caused by the addition of reactive oxygen species 3) analyze the effectiveness of our prevention and treatment.
[1] Horwitz, J. (n.d.). Alpha Crystallin. from http://www.ncbi.nlm.nih.gov/pubmed/12565801
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 87
Illegal BamHI site found at 429 - 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 514
Illegal SapI.rc site found at 374
Sequencing
After running a PCR check and saw the band at the expected size (~1kb), we sent the DNA out for sequencing. The sequencing result is shown below.
- The four restriction sites are highlighted in red
- BBa_K880005 is highlighted in light blue
- CRYAB is highlighted in orange
- 10x Histidine-tag is highlighted in green
- BBa_B0015 is highlighted in dark blue
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