Difference between revisions of "Part:BBa K4308001"
 
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In our project, this primer (<partinfo>BBa_K4308001</partinfo>) was used along with its forward primer (<partinfo>BBa_K4308000</partinfo>) for the amplification reactions of LINC00857, in order to perform isothermal amplification reactions with RT-RPA. Using this method, we were able to detect quantities as low as <var>10<sup>-9</sup></var> nanograms during 30 minutes. The expected amplified product length is about 150 bp.  
 
In our project, this primer (<partinfo>BBa_K4308001</partinfo>) was used along with its forward primer (<partinfo>BBa_K4308000</partinfo>) for the amplification reactions of LINC00857, in order to perform isothermal amplification reactions with RT-RPA. Using this method, we were able to detect quantities as low as <var>10<sup>-9</sup></var> nanograms during 30 minutes. The expected amplified product length is about 150 bp.  
  
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<p style="text-align:center"><b>Figure.1 Schematic diagram of RT-RPA.</b></p>
  
 
===<h1>Characterization</h1>===
 
===<h1>Characterization</h1>===
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<p><b>1.Proof of the expression</b></p>
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<p>After RT-RPA amplification, we carried out 3% agarose gel electrophoresis to verify the amplification results. As can be seen in Figure 1, compared with negative control, our product has obvious bands, which also confirms the validity of the primer.</p>
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<p><b>1. Experimental verification of RT-RPA</b></p>
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<p>The results of the above RPA pre-experiment were better, so we replaced our target gene LINC00857 for reverse transcription-recombinase polymerase amplification (RT-RPA). After RT-RPA amplification, we carried out 3% agarose gel electrophoresis to verify the amplification results. As can be seen in Figure 2, compared with negative control, our product has obvious bands, which also confirms the validity of the primer.</p>
 
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<p style="text-align:center"><b>Figure.1 3% agarose gel electrophoresis to verify the amplification.</b> <i>Lane 1: Target gene (10 nM, centrifuged); Lane 2 : Target gene (1 nM, centrifuged); Lane 3 : Target gene (10 nM, uncentrifuged); Lane 4 : Target gene (1 nM, uncentrifuged). </i></p>
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<p style="text-align:center"><b>Figure.2 3% agarose gel electrophoresis to verify the amplification.</b> <i>Lane 1: Target gene ( 10<sup>1</sup> copies); Lane 2 : Target gene ( 10<sup>2</sup> copies); Lane 3 : Target gene ( 10<sup>3</sup> copies); Lane 4 : Target gene ( 10<sup>4</sup> copies). </i></p>
  
  

Latest revision as of 23:08, 11 October 2022


Reverse Primer for RPA of LINC00857

Long intergenic non-protein coding RNA 857 (LINC00857) is a genetic molecule involved in cancer biology, which is identified as one of the most upregulated LINCRNAs in lung cancer. It could lead to resistance to platinum‐induced apoptosis by affecting mRNA stability or promoting its translation. Thus, we chose LINC00857 as a novel prognostic and cancer predictive biomarker. We use this part as the reverse primer to conduct reverse transcription-recombinase polymerase amplification (RT-RPA) of LINC00857, along with its forward primer (BBa_K4308000).


Usage and Biology

In our project, this primer (BBa_K4308001) was used along with its forward primer (BBa_K4308000) for the amplification reactions of LINC00857, in order to perform isothermal amplification reactions with RT-RPA. Using this method, we were able to detect quantities as low as 10-9 nanograms during 30 minutes. The expected amplified product length is about 150 bp.

Figure.1 Schematic diagram of RT-RPA.

Characterization

1. Experimental verification of RT-RPA

The results of the above RPA pre-experiment were better, so we replaced our target gene LINC00857 for reverse transcription-recombinase polymerase amplification (RT-RPA). After RT-RPA amplification, we carried out 3% agarose gel electrophoresis to verify the amplification results. As can be seen in Figure 2, compared with negative control, our product has obvious bands, which also confirms the validity of the primer.

Figure.2 3% agarose gel electrophoresis to verify the amplification. Lane 1: Target gene ( 101 copies); Lane 2 : Target gene ( 102 copies); Lane 3 : Target gene ( 103 copies); Lane 4 : Target gene ( 104 copies).



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]