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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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Exons and introns of the Cppsy gene in C. protothecoides CS-41.The ten exons are: (1) 1 bp to 280 bp; (2) 477 bp to 576 bp; (3) 691 bp to 743 bp; (4) 913 bp to 1030 bp; (5) 1139 bp to 1180 bp; (6) 1327 bp to 1427 bp; (7) 1635 bp to 1793 bp; (8) 1957 bp to 2045 bp; (9) 2171 bp to 2233 bp; (10) 2351 bp to 2488 bp. (A) Intron density; (B) DNA structure; (C) The relationship between introns and exons of Cppsy, Crpsy, and Cnpsy genes. Dbpsy, Dspsy, Hppsy, Crpsy, Cnpsy, Cppsy, Mzpsy, and Atpsy are the psy genes of Duanliella bardawil, Duanliella salina, Haematococcus pluvialis, Chlamydomonas reinhardtii, Chlorella NC_64A, Chlorella protothecoides CS-41, Zea mays, and Arabidopsis thaliana, respectively.
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marinedrugs-13-06620-f001: Exons and introns of the Cppsy gene in C. protothecoides CS-41.The ten exons are: (1) 1 bp to 280 bp; (2) 477 bp to 576 bp; (3) 691 bp to 743 bp; (4) 913 bp to 1030 bp; (5) 1139 bp to 1180 bp; (6) 1327 bp to 1427 bp; (7) 1635 bp to 1793 bp; (8) 1957 bp to 2045 bp; (9) 2171 bp to 2233 bp; (10) 2351 bp to 2488 bp. (A) Intron density; (B) DNA structure; (C) The relationship between introns and exons of Cppsy, Crpsy, and Cnpsy genes. Dbpsy, Dspsy, Hppsy, Crpsy, Cnpsy, Cppsy, Mzpsy, and Atpsy are the psy genes of Duanliella bardawil, Duanliella salina, Haematococcus pluvialis, Chlamydomonas reinhardtii, Chlorella NC_64A, Chlorella protothecoides CS-41, Zea mays, and Arabidopsis thaliana, respectively.

Mentions: Analysis of the Cppsy gene structure (Figure 1) revealed that it is more complicated than those of dicot and monocot plants. It consists of ten exons and nine introns. Chlorella has a higher intron density than other algae and higher plants; in most of the higher plants, psy genes always have four or five introns, but this alga has nine introns. Compared with the structure of the psy gene from C. reinhardtii (Crpsy), it seems that there are two introns inserted into each of the first and second exons, and one intron inserted into the fourth exon, which makes the gene structure more complicated (Figure 1C).


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)

Exons and introns of the Cppsy gene in C. protothecoides CS-41.The ten exons are: (1) 1 bp to 280 bp; (2) 477 bp to 576 bp; (3) 691 bp to 743 bp; (4) 913 bp to 1030 bp; (5) 1139 bp to 1180 bp; (6) 1327 bp to 1427 bp; (7) 1635 bp to 1793 bp; (8) 1957 bp to 2045 bp; (9) 2171 bp to 2233 bp; (10) 2351 bp to 2488 bp. (A) Intron density; (B) DNA structure; (C) The relationship between introns and exons of Cppsy, Crpsy, and Cnpsy genes. Dbpsy, Dspsy, Hppsy, Crpsy, Cnpsy, Cppsy, Mzpsy, and Atpsy are the psy genes of Duanliella bardawil, Duanliella salina, Haematococcus pluvialis, Chlamydomonas reinhardtii, Chlorella NC_64A, Chlorella protothecoides CS-41, Zea mays, and Arabidopsis thaliana, respectively.
© Copyright Policy
Related In: Results  -  Collection

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

marinedrugs-13-06620-f001: Exons and introns of the Cppsy gene in C. protothecoides CS-41.The ten exons are: (1) 1 bp to 280 bp; (2) 477 bp to 576 bp; (3) 691 bp to 743 bp; (4) 913 bp to 1030 bp; (5) 1139 bp to 1180 bp; (6) 1327 bp to 1427 bp; (7) 1635 bp to 1793 bp; (8) 1957 bp to 2045 bp; (9) 2171 bp to 2233 bp; (10) 2351 bp to 2488 bp. (A) Intron density; (B) DNA structure; (C) The relationship between introns and exons of Cppsy, Crpsy, and Cnpsy genes. Dbpsy, Dspsy, Hppsy, Crpsy, Cnpsy, Cppsy, Mzpsy, and Atpsy are the psy genes of Duanliella bardawil, Duanliella salina, Haematococcus pluvialis, Chlamydomonas reinhardtii, Chlorella NC_64A, Chlorella protothecoides CS-41, Zea mays, and Arabidopsis thaliana, respectively.
Mentions: Analysis of the Cppsy gene structure (Figure 1) revealed that it is more complicated than those of dicot and monocot plants. It consists of ten exons and nine introns. Chlorella has a higher intron density than other algae and higher plants; in most of the higher plants, psy genes always have four or five introns, but this alga has nine introns. Compared with the structure of the psy gene from C. reinhardtii (Crpsy), it seems that there are two introns inserted into each of the first and second exons, and one intron inserted into the fourth exon, which makes the gene structure more complicated (Figure 1C).

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