Difference between revisions of "Team:Aachen/Lab/Methanol/Biobricks"
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− | Before assembling our methanol condensation genes in one genetic construct, we wanted to obtain the required genes separately in a pSB1C3 standard BioBrick vector. | + | Before assembling our methanol condensation genes in one genetic construct, we wanted to obtain the required genes separately in a [http://parts.igem.org/Part:pSB1C3 pSB1C3] standard BioBrick vector. |
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| [http://parts.igem.org/Part:BBa_K1585210 BBa_K1585210] || methanoldehydrogenase 2 || ''mdh'' || 1219 || ''Bacillus methanolicus'' MGA3 | | [http://parts.igem.org/Part:BBa_K1585210 BBa_K1585210] || methanoldehydrogenase 2 || ''mdh'' || 1219 || ''Bacillus methanolicus'' MGA3 | ||
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− | | | + | | [http://parts.igem.org/Part:BBa_K1585211 BBa_K1585211] || 3-hexulose-6-phosphate || ''hps'' || 697 || ''Bacillus methanolicus'' MGA3 |
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− | | | + | | [http://parts.igem.org/Part:BBa_K1585212 BBa_K1585212] || 6-phospho-3-hexuloisomerase || ''phi'' || 616 || ''Bacillus methanolicus'' MGA3 |
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− | | | + | | [http://parts.igem.org/Part:BBa_K1585213 BBa_K1585213] || phosphoketolase || ''xpk'' || 2539 || ''Bifidobacterium adolescentis'' |
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− | Subsequently, we designed and ordered linear gBlocks with additional extensions up- and downstream the coding sequence. Each of the 58 bp upstream extensions contain an overlap to pSB1C3, the BioBrick prefix and the BBa_B0034 ribosome binding site plus the BioBrick scar TAGTAC. Downstream the CDS, we added an 41 bp extension with the suffix and another overlapping region to pSB1C3. | + | Subsequently, we designed and ordered linear gBlocks with additional extensions up- and downstream the coding sequence. Each of the 58 bp upstream extensions contain an overlap to [http://parts.igem.org/Part:pSB1C3 pSB1C3], the BioBrick prefix and the [http://parts.igem.org/Part:BBa_B0034 BBa_B0034] ribosome binding site plus the BioBrick scar TAGTAC. Downstream the CDS, we added an 41 bp extension with the suffix and another overlapping region to [http://parts.igem.org/Part:pSB1C3 pSB1C3]. |
− | Since the CDS of the ''xpk'' is too long for gBlock synthesis, we devided its sequence into two half and designed extensions for both fragments to assemble them together with a linearized pSB1C3 backbone via [[Team:Aachen/Notebook/Protocols#CPEC|CPEC]]. | + | Since the CDS of the ''xpk'' is too long for gBlock synthesis, we devided its sequence into two half and designed extensions for both fragments to assemble them together with a linearized [http://parts.igem.org/Part:pSB1C3 pSB1C3] backbone via [[Team:Aachen/Notebook/Protocols#CPEC|CPEC]]. |
=Results= | =Results= | ||
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− | ''mdh'', ''hps'' and ''phi'' were successfully cloned into the pSB1C3 backbone via restriction and ligation. The two ''xpk'' fragments and a linear pSB1C3 fragment were assembled in a [[Team:Aachen/Notebook/Protocols#CPEC|CPEC]] reaction. All four resulting BioBricks were sequenced and submitted to the BioBrick registry. | + | ''mdh'', ''hps'' and ''phi'' were successfully cloned into the pSB1C3 backbone via restriction and ligation. The two ''xpk'' fragments and a linear [http://parts.igem.org/Part:pSB1C3 pSB1C3] fragment were assembled in a [[Team:Aachen/Notebook/Protocols#CPEC|CPEC]] reaction. All four resulting BioBricks were sequenced and submitted to the BioBrick registry. |
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! BioBrick !! construct !! sequencing confirmed plasmid !! confirmed cryo | ! BioBrick !! construct !! sequencing confirmed plasmid !! confirmed cryo | ||
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− | | | + | | [http://parts.igem.org/Part:BBa_K1585210 BBa_K1585210] || RBS.mdh in [http://parts.igem.org/Part:pSB1C3 pSB1C3] || #9PBD# || #WNFY# |
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− | | | + | | [http://parts.igem.org/Part:BBa_K1585211 BBa_K1585211] || RBS.hps in [http://parts.igem.org/Part:pSB1C3 pSB1C3] || #H4M9# || #863A# |
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− | | | + | | [http://parts.igem.org/Part:BBa_K1585212 BBa_K1585212] || RBS.phi in [http://parts.igem.org/Part:pSB1C3 pSB1C3] || #ENKK# || #AE8V# |
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− | | | + | | [http://parts.igem.org/Part:BBa_K1585213 BBa_K1585213] || RBS.xpk in [http://parts.igem.org/Part:pSB1C3 pSB1C3] || #KFEY# || #8WY9# |
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|} | |} |
Latest revision as of 23:29, 18 September 2015