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Team:Hawaii/Antibiotic test for aadA construct
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== Antibiotic test for ''aadA'' construct == | == Antibiotic test for ''aadA'' construct == | ||
| - | * The gene for Sp and Sm resistance was constructed in BioBrick format. It was amplified from pRL1383a with primers adding BioBrick ends, and the [[Part:BBa_I14032|lac promoter]] and [[Part:BBa_B0034|rbs]] were ligated to it. | + | * The gene for Sp and Sm resistance was constructed in BioBrick format. It was amplified from pRL1383a with primers adding BioBrick ends, and the [[http://partsregistry.org/Part:BBa_I14032| lac promoter]] and [[http://partsregistry.org/Part:BBa_B0034| rbs]] were ligated to it. |
* This construct was tested for antibiotic resistance. | * This construct was tested for antibiotic resistance. | ||
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::*constructs | ::*constructs | ||
::*(+) control pSMC121 (contains ''aadA'' gene) | ::*(+) control pSMC121 (contains ''aadA'' gene) | ||
| - | ::*(-) control pSB1A3 | + | ::*(-) control pSB1A3 |
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== Results == | == Results == | ||
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== Discussion == | == Discussion == | ||
| - | * | + | * AP1 the ''aadA'' gene, the [[http://partsregistry.org/Part:BBa_I14032| the lac promoter]] and an [[http://partsregistry.org/Part:B0034| rbs]] in pSB1A3. This part is similar to [[http://partsregistry.org/Part:BBa_J23012| Spectinomycin R open-reading-frame basic part]] which was built in 2006 and is available in the registry, but the experience states that this part has issues. We checked the sequence of this part and found that it has a point mutation which could be the reason for the lack of expression of this protein. |
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| + | * The [[http://partsregistry.org/Part:BBa_K125530| ''aadA'' gene]] submitted by our team has been verified by sequencing and has been shown to grow on LB+Agar plates containing Sm<sub>50</sub>Sp<sub>100</sub>. | ||
{{Team:Hawaii/Footer}} | {{Team:Hawaii/Footer}} | ||
Latest revision as of 03:19, 29 October 2008
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Contents |
Antibiotic test for aadA construct
- The gene for Sp and Sm resistance was constructed in BioBrick format. It was amplified from pRL1383a with primers adding BioBrick ends, and the lac promoter and rbs were ligated to it.
- This construct was tested for antibiotic resistance.
Methods
- Sequencing confirmation
- Streak on LB+Agar plates with Sm50Sp10
- constructs
- (+) control pSMC121 (contains aadA gene)
- (-) control pSB1A3
Results
- Sequencing confirmation
- Streak
The plate streaked with multiple aadA constructs + controls.
- All four of the constructs grew and the positive control grew while the negative control did not grow.
Discussion
- AP1 the aadA gene, the the lac promoter and an rbs in pSB1A3. This part is similar to Spectinomycin R open-reading-frame basic part which was built in 2006 and is available in the registry, but the experience states that this part has issues. We checked the sequence of this part and found that it has a point mutation which could be the reason for the lack of expression of this protein.
- The aadA gene submitted by our team has been verified by sequencing and has been shown to grow on LB+Agar plates containing Sm50Sp100.
[http://manoa.hawaii.edu/ 
][http://manoa.hawaii.edu/ovcrge/ 
][http://www.ctahr.hawaii.edu 
]
