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Isolation and Analysis of the Cppsy Gene and Promoter from Chlorella protothecoides CS-41.

Li M, Cui Y, Gan Z, Shi C, Shi X - Mar Drugs (2015)

Bottom Line: Analysis revealed several candidate motifs for the promoter, which exhibited light- and methyl jasmonate (MeJA)-responsive characteristics, as well as some typical domains universally discovered in promoter sequences, such as the TATA-box and CAAT-box.Light- and MeJA treatment showed that the Cppsy expression level was significantly enhanced by light and MeJA.These results provide a basis for genetically modifying the carotenoid biosynthesis pathway in C. protothecoides.

View Article: PubMed Central - PubMed

Affiliation: MOST-USDA Joint Research Center for Food Safety, School of Agriculture and Biology, and State Key Lab of Microbial Metabolism, Shanghai Jiao Tong University, Shanghai 200240, China. lmeiya@126.com.

ABSTRACT
Phytoene synthase (PSY) catalyzes the condensation of two molecules of geranylgeranyl pyrophosphate to form phytoene, the first colorless carotene in the carotenoid biosynthesis pathway. So it is regarded as the crucial enzyme for carotenoid production, and has unsurprisingly been involved in genetic engineering studies of carotenoid production. In this study, the psy gene from Chlorella protothecoides CS-41, designated Cppsy, was cloned using rapid amplification of cDNA ends. The full-length DNA was 2488 bp, and the corresponding cDNA was 1143 bp, which encoded 380 amino acids. Computational analysis suggested that this protein belongs to the Isoprenoid_Biosyn_C1 superfamily. It contained the consensus sequence, including three predicted substrate-Mg(2+) binding sites. The Cppsy gene promoter was also cloned and characterized. Analysis revealed several candidate motifs for the promoter, which exhibited light- and methyl jasmonate (MeJA)-responsive characteristics, as well as some typical domains universally discovered in promoter sequences, such as the TATA-box and CAAT-box. Light- and MeJA treatment showed that the Cppsy expression level was significantly enhanced by light and MeJA. These results provide a basis for genetically modifying the carotenoid biosynthesis pathway in C. protothecoides.

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Phylogenetic tree of PSY sequences from various species. The phylogeny was derived using neighbor-joining analysis. The accession numbers of the amino acid sequences follow the taxon names. Horizontal branch lengths represent relative evolutionary distances, with the scale bar corresponding to 0.05 amino acid substitutions per site.
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marinedrugs-13-06620-f002: Phylogenetic tree of PSY sequences from various species. The phylogeny was derived using neighbor-joining analysis. The accession numbers of the amino acid sequences follow the taxon names. Horizontal branch lengths represent relative evolutionary distances, with the scale bar corresponding to 0.05 amino acid substitutions per site.

Mentions: After the DNA and cDNA sequences of the Cppsy gene were determined, it was possible to investigate its evolutionary position among the various psy genes. Using MEGA 4.0 from Clustal W1.6 alignments, the phylogenetic tree of PSYs from different organisms was constructed based on their deduced amino acid sequences. It showed that psy was derived from an ancestor gene and later evolved into four subgroups, including higher plants, cyanobacteria, algae, and bacteria (Figure 2). According to the neighbor-joining (NJ) tree, Cppsy belongs to the algae group, and is more ancient than plant species (Figure 2).


Isolation and Analysis of the Cppsy Gene and Promoter from Chlorella protothecoides CS-41.

Li M, Cui Y, Gan Z, Shi C, Shi X - Mar Drugs (2015)

Phylogenetic tree of PSY sequences from various species. The phylogeny was derived using neighbor-joining analysis. The accession numbers of the amino acid sequences follow the taxon names. Horizontal branch lengths represent relative evolutionary distances, with the scale bar corresponding to 0.05 amino acid substitutions per site.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4663545&req=5

marinedrugs-13-06620-f002: Phylogenetic tree of PSY sequences from various species. The phylogeny was derived using neighbor-joining analysis. The accession numbers of the amino acid sequences follow the taxon names. Horizontal branch lengths represent relative evolutionary distances, with the scale bar corresponding to 0.05 amino acid substitutions per site.
Mentions: After the DNA and cDNA sequences of the Cppsy gene were determined, it was possible to investigate its evolutionary position among the various psy genes. Using MEGA 4.0 from Clustal W1.6 alignments, the phylogenetic tree of PSYs from different organisms was constructed based on their deduced amino acid sequences. It showed that psy was derived from an ancestor gene and later evolved into four subgroups, including higher plants, cyanobacteria, algae, and bacteria (Figure 2). According to the neighbor-joining (NJ) tree, Cppsy belongs to the algae group, and is more ancient than plant species (Figure 2).

Bottom Line: Analysis revealed several candidate motifs for the promoter, which exhibited light- and methyl jasmonate (MeJA)-responsive characteristics, as well as some typical domains universally discovered in promoter sequences, such as the TATA-box and CAAT-box.Light- and MeJA treatment showed that the Cppsy expression level was significantly enhanced by light and MeJA.These results provide a basis for genetically modifying the carotenoid biosynthesis pathway in C. protothecoides.

View Article: PubMed Central - PubMed

Affiliation: MOST-USDA Joint Research Center for Food Safety, School of Agriculture and Biology, and State Key Lab of Microbial Metabolism, Shanghai Jiao Tong University, Shanghai 200240, China. lmeiya@126.com.

ABSTRACT
Phytoene synthase (PSY) catalyzes the condensation of two molecules of geranylgeranyl pyrophosphate to form phytoene, the first colorless carotene in the carotenoid biosynthesis pathway. So it is regarded as the crucial enzyme for carotenoid production, and has unsurprisingly been involved in genetic engineering studies of carotenoid production. In this study, the psy gene from Chlorella protothecoides CS-41, designated Cppsy, was cloned using rapid amplification of cDNA ends. The full-length DNA was 2488 bp, and the corresponding cDNA was 1143 bp, which encoded 380 amino acids. Computational analysis suggested that this protein belongs to the Isoprenoid_Biosyn_C1 superfamily. It contained the consensus sequence, including three predicted substrate-Mg(2+) binding sites. The Cppsy gene promoter was also cloned and characterized. Analysis revealed several candidate motifs for the promoter, which exhibited light- and methyl jasmonate (MeJA)-responsive characteristics, as well as some typical domains universally discovered in promoter sequences, such as the TATA-box and CAAT-box. Light- and MeJA treatment showed that the Cppsy expression level was significantly enhanced by light and MeJA. These results provide a basis for genetically modifying the carotenoid biosynthesis pathway in C. protothecoides.

Show MeSH
Related in: MedlinePlus