Difference between revisions of "Part:BBa K1959000"

(Usage and Biology)
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===Transcriptional activity of ''PSY''===
 
===Transcriptional activity of ''PSY''===
 Semi-quantitative RT-PCR was performed to detect the expression level of ''PSY'' involved in astaxanthin biosynthesis, total RNA of transgenic rice seeds were extracted and cDNA was synthesized from 1μg DNase-treated RNA.
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 Semi-quantitative RT-PCR was performed to detect the expression level of ''PSY'' involved in astaxanthin biosynthesis, total RNA of transgenic rice seeds were extracted and cDNA was synthesized from 1μg DNase-treated RNA. Expected bands of the ''PSY'' gene were observed on the gel, indicated that ''PSY'' gene was transcribed in endosperm.
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[[File:File:T--SCAU-China-PSY-RTPCR.jpg |500px|thumb|centre|<p>'''Figure.2  RT-PCR analyses of expression levels of ''PSY'' genes in several transgenic rice.'''<br>Rice ''OsActin1'' was as an internal control. CK+, positive control (plasmid pYLTAC380MF-BBPC). WT, negative control (wild-type rice cultivar HG1).</p>]]
  
  

Revision as of 17:41, 19 October 2016


Modification of Phytoene synthase (PSY)

The BBa_K1959000 harbors a coding sequence of phytoene synthase (PSY) derived from maize (Zea mays). Codon-optimization has been made for rice (Oryza sativa). PSY catalyzes the conversion of geranylgenranyl diphosphate (GGDP) to phytoene, which is a key enzyme for carotenoid biosynthesis.

Usage and Biology

 Phytoene synthase (PSY, EC 2.5.1.32) is an enzyme involved and responded for the first synthetic reaction in β-carotene biosynthetic pathway. In this reaction, two GGDP are converted to one molecule phytoene and two diphosphates (Figure. 1). PSY genes had been applied to the nutrition-improved crops such as Golden Rice. PSY gene of maize (ZmPSY) was introduced into rice genome to achieve the synthesis of β-carotene. In our project, the ZmPSY was used to reconstruct the pathway of astaxanthin biosynthesis. For maize is a monocots similar to rice, thus the ZmPSY was not fused with transit peptide for plastid sorting.

Figure. 1 Enzymatic reaction catalyzed by PSY

Transcriptional activity of PSY

 Semi-quantitative RT-PCR was performed to detect the expression level of PSY involved in astaxanthin biosynthesis, total RNA of transgenic rice seeds were extracted and cDNA was synthesized from 1μg DNase-treated RNA. Expected bands of the PSY gene were observed on the gel, indicated that PSY gene was transcribed in endosperm.

File:File:T--SCAU-China-PSY-RTPCR.jpg

Figure.2  RT-PCR analyses of expression levels of PSY genes in several transgenic rice.
Rice OsActin1 was as an internal control. CK+, positive control (plasmid pYLTAC380MF-BBPC). WT, negative control (wild-type rice cultivar HG1).


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1045
    Illegal NheI site found at 1164
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 105
    Illegal AgeI site found at 956
  • 1000
    COMPATIBLE WITH RFC[1000]