Difference between revisions of "Part:BBa K747002"
| Line 4: | Line 4: | ||
'''TAL- Proteins''' | '''TAL- Proteins''' | ||
This part can be used to build a 14 nucleotide Transactivator-like (TAL) protein. TAL- Proteins are DNA-targeting modules with repeating Domains, each one of them can bind a specific nucleotide. A plasmid set of 96 DiRepeats (Bba_K747000 to Bba_K747095) allows you to assemble your own specific nucleotide target sequence. Every DiRepeat is able to bind two specific nucleotides and is determined by his position in the 14 repeating Domains. | This part can be used to build a 14 nucleotide Transactivator-like (TAL) protein. TAL- Proteins are DNA-targeting modules with repeating Domains, each one of them can bind a specific nucleotide. A plasmid set of 96 DiRepeats (Bba_K747000 to Bba_K747095) allows you to assemble your own specific nucleotide target sequence. Every DiRepeat is able to bind two specific nucleotides and is determined by his position in the 14 repeating Domains. | ||
| + | |||
| + | synthesis of Transactivator-like (TAL) proteins, i.e. DNA-binding proteins. In contrast to zinc-finger proteins they consist of domain-like repeats in their primary structure that differ only by two amino acids. These two amino acids of a given repeat determine which DNA base the repeat will bind to. By providing efficient mechanisms to combine the different repeats in order to target a certain DNA sequence of around 14bp the FreiGEM team is working towards establishing a standardized kit. | ||
'''Designing a TAL-Protein:''' | '''Designing a TAL-Protein:''' | ||
| − | To build a functional Tal- Protein, you have to choose a 14 nucleotide target sequence. Be aware that the first and the fourteenth nucleotide of the sequence has to be a Thymin. The second to the thirteenth nucleotide can be determined by choosing six DiRepeats of the plasmid set: | + | To build a functional Tal- Protein, you have to choose a 14 nucleotide target sequence. Be aware that the first and the fourteenth nucleotide of the sequence has to be a Thymin. The second to the thirteenth nucleotide can be determined by choosing six DiRepeats of the plasmid set: |
<ul> | <ul> | ||
| Line 15: | Line 17: | ||
<li>Bba_K747064 to Bba_K747079 predefines the nucleotide of the 10. and the 11. nucleotide. | <li>Bba_K747064 to Bba_K747079 predefines the nucleotide of the 10. and the 11. nucleotide. | ||
<li>Bba_K747080 to Bba_K747095 predefines the nucleotide of the 12. and the 13. nucleotide. | <li>Bba_K747080 to Bba_K747095 predefines the nucleotide of the 12. and the 13. nucleotide. | ||
| − | |||
</ul> | </ul> | ||
Revision as of 14:15, 25 September 2012
TAL-Protein_AG1_Direpeat
TAL- Proteins This part can be used to build a 14 nucleotide Transactivator-like (TAL) protein. TAL- Proteins are DNA-targeting modules with repeating Domains, each one of them can bind a specific nucleotide. A plasmid set of 96 DiRepeats (Bba_K747000 to Bba_K747095) allows you to assemble your own specific nucleotide target sequence. Every DiRepeat is able to bind two specific nucleotides and is determined by his position in the 14 repeating Domains.
synthesis of Transactivator-like (TAL) proteins, i.e. DNA-binding proteins. In contrast to zinc-finger proteins they consist of domain-like repeats in their primary structure that differ only by two amino acids. These two amino acids of a given repeat determine which DNA base the repeat will bind to. By providing efficient mechanisms to combine the different repeats in order to target a certain DNA sequence of around 14bp the FreiGEM team is working towards establishing a standardized kit.
Designing a TAL-Protein: To build a functional Tal- Protein, you have to choose a 14 nucleotide target sequence. Be aware that the first and the fourteenth nucleotide of the sequence has to be a Thymin. The second to the thirteenth nucleotide can be determined by choosing six DiRepeats of the plasmid set:
- Bba_K747000 to Bba_K747015 predefines the nucleotide of the 2. and the 3. nucleotide.
- Bba_K747016 to Bba_K747031 predefines the nucleotide of the 4. and the 5. nucleotide.
- Bba_K747032 to Bba_K747047 predefines the nucleotide of the 6. and the 7. nucleotide.
- Bba_K747048 to Bba_K747063 predefines the nucleotide of the 8. and the 9. nucleotide.
- Bba_K747064 to Bba_K747079 predefines the nucleotide of the 10. and the 11. nucleotide.
- Bba_K747080 to Bba_K747095 predefines the nucleotide of the 12. and the 13. nucleotide.
Pipette the following volumes in one tube:
| Component | Volume |
| 6 Direpeats of your choice | 63,3 ng |
| Backbone e. g. TAL-TF | 200 ng ??? |
| BsmBI | 1,5 µl |
| T4 DNA Ligase (60 u) | 1,0 µl |
| T4 DNA Ligase Buffer | 2,0 µl |
| Water, nuclease-free | up to 20 µl |
| Total vloume | 20 µl |
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]