Difference between revisions of "Part:BBa K1959000"
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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. | + | 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.
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.
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
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1045
Illegal NheI site found at 1164 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 105
Illegal AgeI site found at 956 - 1000COMPATIBLE WITH RFC[1000]