IGEM:Imperial/2010/Parts/Registry Upload

Registry Upload

All parts require a basic level of information regarding their sequence, use, design and function. To make information upload as efficient as possible it should all be collated here before submission.

Kirill will provide part numbers!

Necessary information

Copy the following tempalte to ensure that all the required information about each part has been gathered so it can be checked on Monday prior to uploading. An example of how this information should be filled in is included below.

Name: Part name goes here

Code: BBa_K???????

Sequence: ACGT etc.

Short: A short name/decsription of the part (Max. 60 characters inc. spaces)

Long: A thorough description of the part, including what function it has, where it is natively (ie. orignially - eg. E. coli genome locus XYZ) found and what context you have used it in if applicable. References where appropriate!

Source: Where did the part come from (eg. genome, vector). use references where possible!

Design: How was it made? Eg. PCR, Biobrick cloning from X and Y (Use <bbpart></bbpart> function if possible - it doesn't work here but will on the registry). If made by PCr, what enzyme and what primers - e-mail Chris if you don't have their sequences!

References: Very helpful if you have them!

Example

Name: 5’ AmyE Integration Sequence

Code: BBa_K143001

Sequence:

Short: 5’ integration sequence for the AmyE locus of B.subtilis

Long: Integration sequences allow DNA to be incorporated into the chromosome of a host cell at a specific locus using leading (5') and trailing (3') DNA sequences that are the same as those at a specific locus of the chromosome.The 5' integration sequence can be added to the front of a Biobrick construct and the 3' integration sequence specific for this locus (Part BBa_K143002) to the rear of the Biobrick construct to allow integration of the Biobrick construct into the chromosome of the gram positive bacterium B.subtilis.

The AmyE locus was the first locus used for integration into B.subtilis by Shimotsu and Henner[2] and is still commonly used in vectors such as pDR111[3], pDL[4] and their derivatives. Integration at the AmyE locus removes the ability of B.subtilis to break down starch, which can be assayed with iodine as described by Cutting and Vander-horn[5]. The 5' and 3' integration sequences for the AmyE locus were used to integrate the Imperial 2008 iGEM project primary construct into the B.sutbilis chromosome.

Source: The 5’ integration sequence was taken from the shuttle vector pDR111 which has been used in many studies on B.subtilis, in particular in the studies of transcriptional control[3, 6, 7]

Design: The AmyE integration sequence was taken from the vector after comparison by BLAST to the B.subtilis chromosome to identify the homologous sequences. The sequence present in both the host chromosome and the plasmid at the 5' end of the gene is the 5' sequence required for integration

References

2. Shimotsu H and Henner DJ. Construction of a single-copy integration vector and its use in analysis of regulation of the trp operon of Bacillus subtilis. Gene. 1986;43(1-2):85-94. DOI:10.1016/0378-1119(86)90011-9 |
3. Nakano S, Küster-Schöck E, Grossman AD, and Zuber P. Spx-dependent global transcriptional control is induced by thiol-specific oxidative stress in Bacillus subtilis. Proc Natl Acad Sci U S A. 2003 Nov 11;100(23):13603-8. DOI:10.1073/pnas.2235180100 |

4. Bacillus Genetic Stock Center [www.bgsc.org]

5. Cutting, S M.; Vander-Horn, P B. Genetic analysis. In: Harwood C R, Cutting S M. , editors. Molecular biological methods for Bacillus. Chichester, England: John Wiley & Sons, Ltd.; 1990. pp. 27–74.

6. Erwin KN, Nakano S, and Zuber P. Sulfate-dependent repression of genes that function in organosulfur metabolism in Bacillus subtilis requires Spx. J Bacteriol. 2005 Jun;187(12):4042-9. DOI:10.1128/JB.187.12.4042-4049.2005 |
7. Britton RA, Eichenberger P, Gonzalez-Pastor JE, Fawcett P, Monson R, Losick R, and Grossman AD. Genome-wide analysis of the stationary-phase sigma factor (sigma-H) regulon of Bacillus subtilis. J Bacteriol. 2002 Sep;184(17):4881-90. DOI:10.1128/JB.184.17.4881-4890.2002 |

Parts

K316001 - pVEG

Already on Registry

K316002 - dif

Name: B. subtilis dif excision site

Code: BBa_K316002

Sequence:

Short: dif excision site from B. subtilis

Long:

Source: The dif sites were made by annealing synthestised oligoes.

Design: The dif site was made by oligos designed to make overhangs for EcoRI and SpeI ( and ) or XbaI and PstI ( and ) to be used in standard Biobrick or 3A cloning.

References: Very helpful if you have them!