Difference between revisions of "Part:BBa K4012020"
Line 25: Line 25:
 
We construct tSSA1 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 tSSA1(238bp) matched with previous expectations by compared with Marker MK8000 ladder, confirming the successful assembly of Toolkit plasmids and availability in subsequent construction.
 
We construct tSSA1 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 tSSA1(238bp) matched with previous expectations by compared with Marker MK8000 ladder, confirming the successful assembly of Toolkit plasmids and availability in subsequent construction.
  
−
===----- in Level1 plasmid assembly===
+
===tSSA1 in Level1 plasmid assembly===
  
 
[[Image:CDS_DuLAR.jpg|thumbnail|750px|center|'''Figure 2:'''  
 
[[Image:CDS_DuLAR.jpg|thumbnail|750px|center|'''Figure 2:'''  
Line 31: Line 31:
 
The construction schematic of DuLAR sequence demonstrated as Fig.2. The initiation of the DuLAR sequence is done by promoter pPOP6, with termination done by tSSA1. The sequence ConL2 and ConR3 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.2 The band length 2500bp match with expectations. The sequence analysis results show no significant mutations or deletions, representing the success of the assembly.
 
The construction schematic of DuLAR sequence demonstrated as Fig.2. The initiation of the DuLAR sequence is done by promoter pPOP6, with termination done by tSSA1. The sequence ConL2 and ConR3 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.2 The band length 2500bp match with expectations. The sequence analysis results show no significant mutations or deletions, representing the success of the assembly.
  
−
===----- in Level2 plasmid assembly===
+
===tSSA1 in Level2 plasmid assembly===
 
[[Image:yeast PCR.jpg|thumbnail|750px|center|'''Figure 3:'''  
 
[[Image:yeast PCR.jpg|thumbnail|750px|center|'''Figure 3:'''  
 
[https://parts.igem.org/Part:BBa_K4012020] 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]]
 
[https://parts.igem.org/Part:BBa_K4012020] 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]]
 
tSSA1 is also involved in assembly of synthesis pathway of naringenin, shown by Fig.3.
 
tSSA1 is also involved in assembly of synthesis pathway of naringenin, shown by Fig.3.

Revision as of 11:54, 21 October 2021


Terminator.png

tSSA1

  • It is a terminator that ends the sequence that is resposible for the expression of tyrosine ammonia lyase

Secondary Structure

File:Mfold-K4012020-1.png

Measurement

  • [http://openwetware.org/wiki/Cconboy:Terminator_Characterization/Results How these parts were measured]

Obtaining the tSSA1 fragment and BsaI digested verification

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

We construct tSSA1 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 tSSA1(238bp) matched with previous expectations by compared with Marker MK8000 ladder, confirming the successful assembly of Toolkit plasmids and availability in subsequent construction.

tSSA1 in Level1 plasmid assembly

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

The construction schematic of DuLAR sequence demonstrated as Fig.2. The initiation of the DuLAR sequence is done by promoter pPOP6, with termination done by tSSA1. The sequence ConL2 and ConR3 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.2 The band length 2500bp match with expectations. The sequence analysis results show no significant mutations or deletions, representing the success of the assembly.

tSSA1 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

tSSA1 is also involved in assembly of synthesis pathway of naringenin, shown by Fig.3.