Difference between revisions of "Part:BBa K5136039"

 
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<partinfo>BBa_K5136039 short</partinfo>
 
<partinfo>BBa_K5136039 short</partinfo>
  
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===Biology===
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===MntA===
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MntA is a membrane protein from <i>Lactobacillus plantarumATCC 14917</i> which transports Cd<sup>2+</sup> and Mn<sup>2+</sup> into the cell. It is an effective Mn<sup>2+</sup>and Cd<sup>2+</sup> uptake system that exhibits rapid accumulation of the two metal ions. It endows cells with sensitivity to Cd<sup>2+</sup> and energy-dependent Cd<sup>2+</sup> uptake activity. MntA falls into the family of P-type adenosine triphosphatases(ATPase). Its transport efficiency is affected by some factors such as other ion of high concentration and temperature (1,2).
  
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===Usage and Design===
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===Usage and Biology===
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After deinking, heavy metal ions in the ink will be released into the wastewater. Thus, this wastewater needs harmless treatmentremove before discharging. Hence, we use metallothioneins (MTs) to treat wastewater to remove heavy metals. However，due to the poor competitiveness of Cd<sup>2+</sup> in mixed metal ions, engineered bacteria expressing only MTs lack  enough affinity and selectivity for Cd<sup>2+</sup> to reduce Cd<sup>2+</sup> in the wastewater to the levels required by strict government regulations.This basic part (BBa_K5136039) which codes MntA was constructed and then used for the construction of the composite part (<partinfo>BBa_K5136225</partinfo>) .
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===Characterization===
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===Agarose gel electrophoresis (AGE)===
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I0500 promoter was employed to start the expression of MntA (BBa_K5136039) in <I>E. coli</I> DH10β. The basic part (BBa_K5136041) is a component of the composite part (<partinfo>BBa_K5136225</partinfo>). The composite part  (<partinfo>BBa_K5136225</partinfo>) constructed was introduced into the backbone plasmid (pSB1C3) through standard assembly and transformed into <I>E. coli</I> DH10β. The positive clones were selected, and colony PCR and gene sequencing were used to verify that the clones were correct . Target bands (2584 bp) can be observed at the position between 2000bp and 3000bp (Figure 1).
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<center><html><img src="https://static.igem.wiki/teams/5136/part/kyh/2251.png"width="200px"></html></center>
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<center><b>Figure 1 Colony PCR of BBa_K5136225_pSB3K3 in <i>E. coli</i> BL21(DE3)</b></center>
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===Reference===
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1. Z. Hao, S. Chen, D. B. Wilson, Cloning, Expression, and Characterization of Cadmium and Manganese Uptake Genes from Lactobacillus Plantarum. Appl Environ Microbiol 65, 4746-4752 (1999).
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2. S. K. Kim, B. S. Lee, D. B. Wilson, E. K. Kim, Selective Cadmium Accumulation Using Recombinant Escherichia Coli. J Biosci Bioeng 99, 109-114 (2005).
  
 
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Revision as of 17:08, 1 October 2024


mnta

Biology

MntA

MntA is a membrane protein from Lactobacillus plantarumATCC 14917 which transports Cd2+ and Mn2+ into the cell. It is an effective Mn2+and Cd2+ uptake system that exhibits rapid accumulation of the two metal ions. It endows cells with sensitivity to Cd2+ and energy-dependent Cd2+ uptake activity. MntA falls into the family of P-type adenosine triphosphatases(ATPase). Its transport efficiency is affected by some factors such as other ion of high concentration and temperature (1,2).

Usage and Design

After deinking, heavy metal ions in the ink will be released into the wastewater. Thus, this wastewater needs harmless treatmentremove before discharging. Hence, we use metallothioneins (MTs) to treat wastewater to remove heavy metals. However，due to the poor competitiveness of Cd2+ in mixed metal ions, engineered bacteria expressing only MTs lack enough affinity and selectivity for Cd2+ to reduce Cd2+ in the wastewater to the levels required by strict government regulations.This basic part (BBa_K5136039) which codes MntA was constructed and then used for the construction of the composite part (BBa_K5136225) .

Characterization

Agarose gel electrophoresis (AGE)

I0500 promoter was employed to start the expression of MntA (BBa_K5136039) in E. coli DH10β. The basic part (BBa_K5136041) is a component of the composite part (BBa_K5136225). The composite part  (BBa_K5136225) constructed was introduced into the backbone plasmid (pSB1C3) through standard assembly and transformed into E. coli DH10β. The positive clones were selected, and colony PCR and gene sequencing were used to verify that the clones were correct . Target bands (2584 bp) can be observed at the position between 2000bp and 3000bp (Figure 1).

Figure 1 Colony PCR of BBa_K5136225_pSB3K3 in E. coli BL21(DE3)

Reference

1. Z. Hao, S. Chen, D. B. Wilson, Cloning, Expression, and Characterization of Cadmium and Manganese Uptake Genes from Lactobacillus Plantarum. Appl Environ Microbiol 65, 4746-4752 (1999).
2. S. K. Kim, B. S. Lee, D. B. Wilson, E. K. Kim, Selective Cadmium Accumulation Using Recombinant Escherichia Coli. J Biosci Bioeng 99, 109-114 (2005).

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 128
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 436
    Illegal AgeI site found at 1028
    Illegal AgeI site found at 1537
  • 1000
    COMPATIBLE WITH RFC[1000]