Team:Technion/Attributions

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Revision as of 22:07, 26 September 2012

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Attributions

During our project we have used parts that were given to us by different labs or were taken from the registry/distribution kit.

Donated parts

• Ann K. Ganesan - WT T7 RNAP
• Changwon Kang - WT SP6 RNAP
• Christopher A. Voight - T7*, T7*(T3), T7*(N4), T7*(K1F) RNAPs with suitable induced promoters: pT7, pT3, pK1F, pN4
• Gallivan Lab (Gallivan Justin P., Lynch Sean A.) - Theophylline riboswitch and pTAC
• Roee's Lab (Roee Amit) - Cerulean FP under control of pLac/Ara, mCitrin FP and mCherry under control of pTetO.

Parts from Distribution Kit

General

• The image of the bacteriophage used for the creation of our banner was made by Jonathan Heras from Equinox Graphics Ltd. and was found on science magazine 2010 visualization challenge

Achievements

• We have submitted DNA for 25 BioBricks to the registry. Due to lack of time we haven't managed to put all the parts inside the shipping plasmid.

• We have created new reporter system to test our YES gates and hope that it will become standard system for testing other parts as well. We used it to characterize T7 RNAP activity, the results can be seen here.

• We have managed to successfully clone 4 out of 8 lambda phage fragments that the phage's genome has been divided into.

• We characterized a theophylline riboswitch+mCherry construct. The results are here.

• We have managed to combine a theophylline riboswitch with T7, T7*(T3), T7*(K1F), T7*(N4) by using fusion PCR under the control of a tac promoter, more details can be seen at RS section. The riboswitch with T7 and T7*(N4) were cloned into the shipping plasmid and submitted as BioBricks. Due to lack of time we hadn't transferred RS+RNAP constructs under pLux control, as was planned at the beginning.

• We have produced a working YES gate #1 with T7* RNAP. It consisted of a P_TetO promoter followed by mCherry, T7* RNAP and a terminator.

• We have added MCS to pSB1C3 plasmid backbone. This backbone is very convenient to work with, some of our parts had the same restriction sites as standard prefix and suffix found in the backbone, so we couldn't insert them into the pSB1C3 plasmid. Addition of MCS helps to circumvent this problem.

• We entered experience for the following pre-existing BioBricks: pSB3C5 and BBa_I0462.

• Some of our parts (particularly the phage fragments) were incompatible with current restriction based registry standards. Therefore, we contacted the registry, asking them to let us send parts that are incompatible with the standards. The registry told us that we must obey some registry standard or create a new standard. Thus, we decided to create a BioBrick standard to be used with Gibson Assembly, releasing users from the restriction sites limitations. Since this idea was conceived only a couple of days before the deadline, we didn't manage to develop it fully. Briefly, it can be suggested that a prefix and a suffix for a standard backbone would not include any restriction sites. Those prefix and suffix will be used for Gibson assembly of BioBricks into that backbone. For assembly of two BioBricks or more into the same backbone, the method suggested by the Gibson assembly manual may be used along with the standardized prefix and suffix.