Difference between revisions of "Part:BBa K4012008"

 
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<partinfo>BBa_K4012008 parameters</partinfo>
 
<partinfo>BBa_K4012008 parameters</partinfo>
 
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===Obtaining the pAOX1 fragment and BsaI digested verification===
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[[Image:T--AISSU_Union--partslast.jpg|thumbnail|750px|center|'''Figure 1:'''
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[https://parts.igem.org/Part:BBa_K4012008] Results of yeast toolkit plasmids enzyme-digested verification]]
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We construct pAOX1 with vector type8-pSB1K3-GFP and proceed digested verification using BsaI. According to Fig.1, we obtain two clear bands after BsaI digested, the length of the vector(1622bp) and the inserted fragments pAOX1(943bp) matched with previous expectations by compared with Marker MK8000 ladder, confirming the successful assembly of Toolkit plasmids and availability in subsequent construction.
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===pAOX1 in Level1 plasmid assembly===
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[[Image:CPR.jpg|thumbnail|750px|center|'''Figure 2:'''
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[https://parts.igem.org/Part:BBa_K4012008] A: the construction strategy; B: results of colony PCR; C: results of sequencing analysis of coding sequence MsCHI]]
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The construction schematic of MsCHI sequence demonstrated as Fig .6. The initiation of the MsCHI sequence is done by promoter pAOX1, with termination done by tTDH1. The sequence ConL3 and ConRE are connector sequences within the Level 1 plasmid assembly. Furthermore, typr9-KVF and type9-VR are imposed to enact selection through colonies PCR, results shown in Fig.6 The band length 2000bp match with expectations. The sequence analysis results show no significant mutations or deletions, representing the success of the assembly.
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===pAOX1 in Level2 plasmid assembly===
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[[Image:yeast PCR.jpg|thumbnail|750px|center|'''Figure 3:'''
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[https://parts.igem.org/Part:BBa_K4012008] A: The results of colony PCR of upstream transformant; B: The results of colony PCR of downstream transformant; C: Construction of catechin metabolic pathway; D：Construction of naringenin metabolic pathway]]
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pAOX1 is also involved in assembly of synthesis pathway of naringenin, shown by Fig.3.

Revision as of 11:37, 21 October 2021


pAOX1

it is a promotor that starts the sequence that is resposible for the production of chalcone isomerase

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Obtaining the pAOX1 fragment and BsaI digested verification

Figure 1: [1] Results of yeast toolkit plasmids enzyme-digested verification

We construct pAOX1 with vector type8-pSB1K3-GFP and proceed digested verification using BsaI. According to Fig.1, we obtain two clear bands after BsaI digested, the length of the vector(1622bp) and the inserted fragments pAOX1(943bp) matched with previous expectations by compared with Marker MK8000 ladder, confirming the successful assembly of Toolkit plasmids and availability in subsequent construction.

pAOX1 in Level1 plasmid assembly

Figure 2: [2] A: the construction strategy; B: results of colony PCR; C: results of sequencing analysis of coding sequence MsCHI

The construction schematic of MsCHI sequence demonstrated as Fig .6. The initiation of the MsCHI sequence is done by promoter pAOX1, with termination done by tTDH1. The sequence ConL3 and ConRE are connector sequences within the Level 1 plasmid assembly. Furthermore, typr9-KVF and type9-VR are imposed to enact selection through colonies PCR, results shown in Fig.6 The band length 2000bp match with expectations. The sequence analysis results show no significant mutations or deletions, representing the success of the assembly.

pAOX1 in Level2 plasmid assembly

Figure 3: [3] A: The results of colony PCR of upstream transformant; B: The results of colony PCR of downstream transformant; C: Construction of catechin metabolic pathway; D：Construction of naringenin metabolic pathway

pAOX1 is also involved in assembly of synthesis pathway of naringenin, shown by Fig.3.