Difference between revisions of "Part:BBa K343004"
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<partinfo>BBa_K343004 short</partinfo>
 
<partinfo>BBa_K343004 short</partinfo>
  
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<p style="text-align: justify;">
 
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<br>
 
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This part entails a TetR repressed POPS/RIPS generator (including RBS), the FlhDC master operon and a dual-terminator. This part will cause hyperflagellated cells.
 
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<br>
 
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</p>
 
  
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== Part background ==
 
  
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<p style="text-align: justify;">
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=Beta-carotene monooxygenase=
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<br>
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The flagella regulon in ''E. coli'' is composed of at least 50 genes organized in no less than 14 operons that all contribute to the synthesis and operation of flagella. The operons are synthesized in a three-level transcriptional cascade where the ''FlhDC'' operon is the master regulator at the top of the cascade. The flagella regulon is tightly controlled by nutritional and environmental conditions, ''E. coli'' starved of amino acids showed temporarily decrease of the flagella regulon transcripts which are needed for the synthesis and operation of the flagellum.[http://onlinelibrary.wiley.com/doi/10.1111/j.1365-2958.2009.06939.x/full (1)]
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The synthesis and assembly of flagella are regulated by the transcriptional cascade composed of three levels of gene products (class I, -II and –III). Class I genes consist of a single operon encoding the proteins ''FlhD'' and ''FlhC'' that form a multimeric (''FlhD4C2'') transcriptional activation complex. This ‘master regulator’ stimulates transcription by binding upstream of Class II promoters. Class II genes encode proteins that assemble to form the basal body and hook of the flagellum, as well as the ''fliA'' gene that encodes the alternative σ factor σ28, also called σF. σ28 binds to RNA polymerase (RNAP) core enzyme and directs it to Class III promoters. Class III genes encode the rest of the structural genes of the flagellum, including ''fliC'' encoding flagellin, as well as the chemotaxis apparatus. [http://onlinelibrary.wiley.com/doi/10.1111/j.1365-2958.2009.06939.x/full (1)]
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==Background==
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<br>
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It has been shown that overexpression of the ''FlhDC'' operon restores motility in mutants that have been made immotile [http://jb.asm.org/cgi/content/short/181/24/7500 (2)]. Also, overexpression of ''FlhDC'' in the ''E. coli'' K12 strain MG1655 made the cells hypermotile.[http://iai.asm.org/cgi/content/abstract/75/7/3315 (3)]
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<br>
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===Beta-carotene monooxygenase gene===
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</p>
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 +
====Protein structure====
 +
 
 +
 
 +
===Retinal===
 +
 
 +
==Usage and parameters==
 +
 
 +
===Usage===
 +
 
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 +
===Performance===
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'''Response time''':
 +
'''Production rate''':  
 +
 
 +
'''Plasmid stability''':
 +
 
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'''Growth rate''':
 +
 
 +
===Compatibility===
 +
This brick has been tested in the following plasmids and stains:
 +
 
 +
'''Chassis''': ''E. coli'' TOP10, ''E. coli'' MG1655.  
 +
 
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'''Plasmids''': [https://parts.igem.org/Part:pSB1C3 PSB1C3] (high-copy), [https://parts.igem.org/Part:pSB3C5 PSB3C5] (low-copy).
 +
 
 +
'''Devices''': Device has been shown to work with [https://parts.igem.org/Part:BBa_K274210 BBa_K274210].
 +
 
 +
===Safety===
 +
'''General use''': It is our general consensus that this BioBrick does not pose any treat to trained peopled working in a level 1 lab. No special care is needed when working with the BioBrick.
 +
 
 +
'''Potential pathogenicity''': We do not recommend using this BioBrick for any type of system in humans or animals.
 +
 
 +
'''Environmental impact''': This BioBrick can be used under controlled settings, but not recommended in the wild.  
 +
 
 +
Please see our risk assessment as to why we came to these conclusions.
 +
 
 +
==Risk-assessment==
 +
 
 +
====General use====
 +
 
 +
<p style="text-align: justify;">This BioBrick poses no treat to the welfare of people working with it, as long as this is done in at least a level 1 safety lab by trained people. No special care is needed when working with this BioBrick. </p>
 +
 
 +
====Potential pathogenicity====
 +
 
 +
<p style="text-align: justify;">This BioBrick consists of three different parts: The first 224 amino acid residues come from the ''NpSopII'' gene from ''Natronomonas pharaonis'', encoding a blue-light photon receptor with 15 residues removed at the C-terminal. The following 9 amino acids are a linker. The last part is ''HtrII'' fused with ''Tar'' from ''E. coli''. The complex' first 125 amino acid residues come from ''HtrII'' and the remaining 279 from ''Tar'' ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 7]). ''NpHtrII'' is thought to function in signal transduction and activation of microbial signalling cascades ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 8]).  </p>
 +
 
 +
<p style="text-align: justify;">A single article has been written about haloarchaea in humans indicating that these played a role in patients with inflammatory bowel disease ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 9]), but there is no evidence that the genes this BioBrick is made from or any near homologs are involved in any disease processes, toxic products or invasion properties. They do not regulate the immune system in any way.</p>
 +
 
 +
====Environmental impact====
 +
 
 +
<p style="text-align: justify;">The BioBrick does not produce a product that is secreted into the environment, nor is it’s gene product itself toxic. It would not produce anything that distrupt natural occurring symbiosis.</p>
 +
 
 +
<p style="text-align: justify;">The BioBrick might increase a bacteria’s ability to find nutrients and as such ease its ability to replicate and spread in certain dark environments. On the other hand the BioBrick is very large and this will naturally slow down its replication rate. Generally we do not believe this BioBrick will make its host able to outcompete natural occurring bacteria, simply because it’s function is not something that will give its host a functional advantage.  </p>
 +
 
 +
====Possible malign use====
 +
 
 +
<p style="text-align: justify;">This BioBrick will not increase its hosts ability to survive in storage conditions, to be aerosoled, to be vaporized or create spores. None of its proteins regulate or affect the immune system or are pathogenic towards humans and animals.</p>==Resources==
 +
 
 +
Datasheet for BioBrick.
 +
 
 +
PDB file for protein structure.
 +
 
 +
==References==
 +
 
 +
#ENZYME entry 1.14.99.36 [Internet]. [cited 2010 Oct 13];Available from: http://www.expasy.org/cgi-bin/nicezyme.pl?1.14.99.36
 +
#von Lintig J, Dreher A, Kiefer C, Wernet MF, Vogt K. Analysis of the blind Drosophila mutant ninaB identifies the gene encoding the key enzyme for vitamin A formation in vivo. Proceedings of the National Academy of Sciences of the United States of America. 2001 Jan 30;98(3):1130 -1135.
 +
#Retinal - Wikipedia, the free encyclopedia [Internet]. [cited 2010 Oct 13];Available from: http://en.wikipedia.org/wiki/Retinal
 +
#Part:BBa K274210 - parts.igem.org [Internet].  [cited 2010 Oct 13];Available from: https://parts.igem.org/Part:BBa_K274210
 +
#Bryant DA, Frigaard N. Prokaryotic photosynthesis and phototrophy illuminated. Trends Microbiol. 2006 Nov;14(11):488-496.
 +
#Retinaldehyde - PubChem Public Chemical Database [Internet].  [cited 2010 Oct 13];Available from: http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=1070
 +
#ninaB neither inactivation nor afterpotential B [Drosophila melanogaster] - Gene result [Internet].  [cited 2010 Oct 13];Available from: http://www.ncbi.nlm.nih.gov/gene/41678
 +
#von Lintig J, Vogt K. Filling the Gap in Vitamin A Research. Journal of Biological Chemistry. 2000 Apr 21;275(16):11915 -11920.
 +
#ENZYME: 1.14.99.36 [Internet].  [cited 2010 Oct 13];Available from:http://www.genome.jp/dbget-bin/www_bget?ec:1.14.99.36
 +
#Kelley LA & Sternberg MJE. Protein structure prediction on the web: a case study using the Phyre server. Nature Protocols. 4, 363 - 371 (2009).
 +
#Spiegl N, Didichenko S, McCaffery P, Langen H, Dahinden CA. Human basophils activated by mast cell-derived IL-3 express retinaldehyde dehydrogenase-II and produce the immunoregulatory mediator retinoic acid. Blood. 2008 Nov 1;112(9):3762-71.
 +
#Russell RM. The vitamin A spectrum: from deficiency to toxicity. American Journal of Clinical Nutrition, Vol. 71, No. 4, 878-884, April 2000.
 +
#Pasquali D, Thaller C, Eichele G. Abnormal level of retinoic acid in prostate cancer tissues. J Clin Endocrinol Metab. 1996 Jun;81(6):2186-91.
 +
 
 +
 
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K343004 SequenceAndFeatures</partinfo>
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<partinfo>BBa_K343006 SequenceAndFeatures</partinfo>
  
  
 
<!-- Uncomment this to enable Functional Parameter display  
 
<!-- Uncomment this to enable Functional Parameter display  
 
===Functional Parameters===
 
===Functional Parameters===
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<partinfo>BBa_K343004 parameters</partinfo>
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<partinfo>BBa_K343006 parameters</partinfo>
 
<!-- -->
 
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Revision as of 19:45, 27 October 2010

Flagella overekspression


Beta-carotene monooxygenase

Background

Beta-carotene monooxygenase gene

Protein structure

Retinal

Usage and parameters

Usage

Performance

Response time: Production rate:

Plasmid stability:

Growth rate:

Compatibility

This brick has been tested in the following plasmids and stains:

Chassis: E. coli TOP10, E. coli MG1655.

Plasmids: PSB1C3 (high-copy), PSB3C5 (low-copy).

Devices: Device has been shown to work with BBa_K274210.

Safety

General use: It is our general consensus that this BioBrick does not pose any treat to trained peopled working in a level 1 lab. No special care is needed when working with the BioBrick.

Potential pathogenicity: We do not recommend using this BioBrick for any type of system in humans or animals.

Environmental impact: This BioBrick can be used under controlled settings, but not recommended in the wild.

Please see our risk assessment as to why we came to these conclusions.

Risk-assessment

General use

This BioBrick poses no treat to the welfare of people working with it, as long as this is done in at least a level 1 safety lab by trained people. No special care is needed when working with this BioBrick.

Potential pathogenicity

This BioBrick consists of three different parts: The first 224 amino acid residues come from the NpSopII gene from Natronomonas pharaonis, encoding a blue-light photon receptor with 15 residues removed at the C-terminal. The following 9 amino acids are a linker. The last part is HtrII fused with Tar from E. coli. The complex' first 125 amino acid residues come from HtrII and the remaining 279 from Tar ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 7]). NpHtrII is thought to function in signal transduction and activation of microbial signalling cascades ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 8]).

A single article has been written about haloarchaea in humans indicating that these played a role in patients with inflammatory bowel disease ([http://2010.igem.org/Team:SDU-Denmark/safety-b#References 9]), but there is no evidence that the genes this BioBrick is made from or any near homologs are involved in any disease processes, toxic products or invasion properties. They do not regulate the immune system in any way.

Environmental impact

The BioBrick does not produce a product that is secreted into the environment, nor is it’s gene product itself toxic. It would not produce anything that distrupt natural occurring symbiosis.

The BioBrick might increase a bacteria’s ability to find nutrients and as such ease its ability to replicate and spread in certain dark environments. On the other hand the BioBrick is very large and this will naturally slow down its replication rate. Generally we do not believe this BioBrick will make its host able to outcompete natural occurring bacteria, simply because it’s function is not something that will give its host a functional advantage.

Possible malign use

This BioBrick will not increase its hosts ability to survive in storage conditions, to be aerosoled, to be vaporized or create spores. None of its proteins regulate or affect the immune system or are pathogenic towards humans and animals.

==Resources==

Datasheet for BioBrick.

PDB file for protein structure.

References

  1. ENZYME entry 1.14.99.36 [Internet]. [cited 2010 Oct 13];Available from: http://www.expasy.org/cgi-bin/nicezyme.pl?1.14.99.36
  2. von Lintig J, Dreher A, Kiefer C, Wernet MF, Vogt K. Analysis of the blind Drosophila mutant ninaB identifies the gene encoding the key enzyme for vitamin A formation in vivo. Proceedings of the National Academy of Sciences of the United States of America. 2001 Jan 30;98(3):1130 -1135.
  3. Retinal - Wikipedia, the free encyclopedia [Internet]. [cited 2010 Oct 13];Available from: http://en.wikipedia.org/wiki/Retinal
  4. Part:BBa K274210 - parts.igem.org [Internet]. [cited 2010 Oct 13];Available from: https://parts.igem.org/Part:BBa_K274210
  5. Bryant DA, Frigaard N. Prokaryotic photosynthesis and phototrophy illuminated. Trends Microbiol. 2006 Nov;14(11):488-496.
  6. Retinaldehyde - PubChem Public Chemical Database [Internet]. [cited 2010 Oct 13];Available from: http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=1070
  7. ninaB neither inactivation nor afterpotential B [Drosophila melanogaster] - Gene result [Internet]. [cited 2010 Oct 13];Available from: http://www.ncbi.nlm.nih.gov/gene/41678
  8. von Lintig J, Vogt K. Filling the Gap in Vitamin A Research. Journal of Biological Chemistry. 2000 Apr 21;275(16):11915 -11920.
  9. ENZYME: 1.14.99.36 [Internet]. [cited 2010 Oct 13];Available from:http://www.genome.jp/dbget-bin/www_bget?ec:1.14.99.36
  10. Kelley LA & Sternberg MJE. Protein structure prediction on the web: a case study using the Phyre server. Nature Protocols. 4, 363 - 371 (2009).
  11. Spiegl N, Didichenko S, McCaffery P, Langen H, Dahinden CA. Human basophils activated by mast cell-derived IL-3 express retinaldehyde dehydrogenase-II and produce the immunoregulatory mediator retinoic acid. Blood. 2008 Nov 1;112(9):3762-71.
  12. Russell RM. The vitamin A spectrum: from deficiency to toxicity. American Journal of Clinical Nutrition, Vol. 71, No. 4, 878-884, April 2000.
  13. Pasquali D, Thaller C, Eichele G. Abnormal level of retinoic acid in prostate cancer tissues. J Clin Endocrinol Metab. 1996 Jun;81(6):2186-91.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 765
    Illegal BamHI site found at 500
    Illegal BamHI site found at 1757
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1361
    Illegal BsaI site found at 1809