Coding

Part:BBa_K3867005

Designed by: Lei Huang   Group: iGEM21_ICJFLS   (2021-10-13)
Revision as of 11:35, 14 October 2021 by Leo Huang (Talk | contribs)

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APL1 (ADP glucose pyrophosphorylase large subunit 1)

ADP glucose pyrophosphorylase, abbreviated as AGPase, containing 2 subunits, ADG1 and APL1. ADG1 encodes the small subunit and APL1 encodes the large subunit. The large subunit plays a regulatory role whereas the small subunit (ADG1) is the catalytic isoform responsible for AGPase activity. AGPase catalyzes Glucose-6-phosphate (Glu-6-P) converted to adenosine diphosphate glucose (ADPG), which is a key enzyme in amylose synthesis and catalyzes the first, rate limiting step in amylose biosynthesis.

K3867001-1.jpg


APL1 gene was cloned into the vector pSB1C3. PCR method was used to identify. Fig.1 shows the result of identification.

K3867002-2.jpg

Fig.1. The result of APL1 gene cloning. M: Marker; 1: plasmid of pSB1C3-APL1; 2: PCR result of APL1 gene.

When APL1 gene was constructed to the expression vector pETDuet-1, APL1 protein was expressed. His tag was used to purify APL1 protein. The identification result was showed in Fig.2

K3867002-3.jpg



Fig.2. The expression of APL1 with vector pETDuet-1-APL1 in E.coli. M, Marker; 1, No induced by IPTG; 2, Purification of APL1 protein (57.5kD) induced by IPTG; 3, induced by IPTG.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1069
    Illegal BglII site found at 1510
    Illegal BamHI site found at 255
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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