Difference between revisions of "Part:BBa K4083000"
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<partinfo>BBa_K4083000 short</partinfo>
 
<partinfo>BBa_K4083000 short</partinfo>
  
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Duo2 plasmid is engineered by Rahul Gauttam, Aindrila Mukhopadhyay, and Steven W. Singer (2020)[1]. The plasmid was sent to us by Rahul Gauttam who is currently working as a postdoctoral fellow at Berkeley Lab.  
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Duo2 plasmid is engineered by Rahul Gauttam, Aindrila Mukhopadhyay, and Steven W. Singer (2020)[1]. The plasmid was sent to us by Rahul Gauttam who is currently working as a postdoctoral fellow at Berkeley Lab. This plasmid was modified from the pRG_Duet vector for Corynebacterium glutamicum.  
  
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About plasmid
 
  
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This plasmid was modified from the pRG_Duet vector for Corynebacterium glutamicum.
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===Usage and Biology===
  
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Reference:
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https://parts.igem.org/Part:BBa_K4083000:Design
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Figure 1. Illustration of Duo2 lasmid.
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As illustrated in Figure 1, this plasmid contains two multiple cloning sites (MCS 1 and MCS2) with two transcriptional terminators. The MCS1 and MCS2 are regulated by IPTG-inducible Ptac and aTc-inducible PtetR/tetA promoters respectively. Moreover, there are lacI and tetR dependent repression systems that are regulated by Ptac and PtetR/tetA respectively. They were integrated for metabolic engineering purposes. There is a kanamycin resistance gene (KanR). There are two origins of replication to allow replication in E. coli (colE1) and P. putida (pRO1600).
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Our team incorporated the rhla/rhlb and nadE genes into MCS1 and MCS2 respectively. Our team planned to extract rhL and rhlB genes from P.aeruginosa and to insert them into pRGPDuo2 plasmid. We developed the new approach to increase the P. putida's rhamnolpid synthesis by adding nadE gene which encodes NAD synthetase. This way, we hoped to see more rhamnolipid production in engineered P. putida.
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===Part functionality===
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===Reference===
  
 
[1] Gauttam, R., Mukhopadhyay, A., & Singer, S. W. (2020). Construction of a novel dual-inducible duet-expression system for gene (over)expression in Pseudomonas putida. Plasmid, 110. https://doi.org/10.1016/j.plasmid.2020.102514
 
[1] Gauttam, R., Mukhopadhyay, A., & Singer, S. W. (2020). Construction of a novel dual-inducible duet-expression system for gene (over)expression in Pseudomonas putida. Plasmid, 110. https://doi.org/10.1016/j.plasmid.2020.102514

Revision as of 10:06, 20 October 2021


pRGPDuo2 plasmid for P. putida

Duo2 plasmid is engineered by Rahul Gauttam, Aindrila Mukhopadhyay, and Steven W. Singer (2020)[1]. The plasmid was sent to us by Rahul Gauttam who is currently working as a postdoctoral fellow at Berkeley Lab. This plasmid was modified from the pRG_Duet vector for Corynebacterium glutamicum.


Usage and Biology

https://parts.igem.org/Part:BBa_K4083000:Design

Figure 1. Illustration of Duo2 lasmid.

As illustrated in Figure 1, this plasmid contains two multiple cloning sites (MCS 1 and MCS2) with two transcriptional terminators. The MCS1 and MCS2 are regulated by IPTG-inducible Ptac and aTc-inducible PtetR/tetA promoters respectively. Moreover, there are lacI and tetR dependent repression systems that are regulated by Ptac and PtetR/tetA respectively. They were integrated for metabolic engineering purposes. There is a kanamycin resistance gene (KanR). There are two origins of replication to allow replication in E. coli (colE1) and P. putida (pRO1600).

Our team incorporated the rhla/rhlb and nadE genes into MCS1 and MCS2 respectively. Our team planned to extract rhL and rhlB genes from P.aeruginosa and to insert them into pRGPDuo2 plasmid. We developed the new approach to increase the P. putida's rhamnolpid synthesis by adding nadE gene which encodes NAD synthetase. This way, we hoped to see more rhamnolipid production in engineered P. putida.

Part functionality

Reference

[1] Gauttam, R., Mukhopadhyay, A., & Singer, S. W. (2020). Construction of a novel dual-inducible duet-expression system for gene (over)expression in Pseudomonas putida. Plasmid, 110. https://doi.org/10.1016/j.plasmid.2020.102514

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 3853
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 3880
    Illegal BamHI site found at 31
    Illegal XhoI site found at 2175
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 6480
  • 1000
    COMPATIBLE WITH RFC[1000]