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DNA fingerprinting differentiation between beta-carotene hyperproducer strains of Dunaliella from around the world.

Olmos J, Ochoa L, Paniagua-Michel J, Contreras R - Saline Syst. (2009)

Bottom Line: In this work, we applied our intron-sizing method to compare the 18S rDNA fingerprint between D. salina (CCAP 19/18), D. salina/bardawil (UTEX LB2538) and beta-carotene hyperproducing strains of Dunaliella isolated from salt saturated lagoons in Baja, Mexico.In Baja Mexico we found D. salina and D. salina/bardawil species by using intron-sizing-method.The National Center for Biotechnology Information (NCBI) Dunaliella 18S rDNA gene sequences were analyzed with our methodology and extraordinary correlation was found with experimental results.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Microbiology Laboratory, Centro de Investigación Científica y de Educación Superior de Ensenada (CICESE), Department of Marine Biotechnology, Ensenada, B.C, México.

ABSTRACT

Background: Dunaliella salina is the most important species of the genus for beta-carotene production. Several investigations have demonstrated that D. salina produces more than 10% dry weight of pigment and that the species grows in salt saturated lagoons. High plasticity in the green stage and the almost indistinguishable differences in the red phase make identification and differentiation of species and ecotypes very difficult and time consuming.

Results: In this work, we applied our intron-sizing method to compare the 18S rDNA fingerprint between D. salina (CCAP 19/18), D. salina/bardawil (UTEX LB2538) and beta-carotene hyperproducing strains of Dunaliella isolated from salt saturated lagoons in Baja, Mexico. All hyperproducer strains reached beta-carotene levels of about 10 pg/cell. Optical microscopy did not allow to differentiate between these Dunaliella strains; however, 18S rDNA fingerprinting methodology allowed us to differentiate D. salina from D. salina/bardawil.

Conclusion: In Baja Mexico we found D. salina and D. salina/bardawil species by using intron-sizing-method. The National Center for Biotechnology Information (NCBI) Dunaliella 18S rDNA gene sequences were analyzed with our methodology and extraordinary correlation was found with experimental results.

No MeSH data available.


Amplification with conserved and specific oligonucleotides. Lane 1, molecular weight marker. Lane 2 and 3 corresponds to amplification with MA1–MA2 conserved primers, using DNA samples from San Quintín and La Salina, respectively. Lane 4 and 5, corresponds to amplification with DBs-MA2 and DSs-MA2 specific-conserved oligonucleotides, using both β-carotene hyperproducers strains.
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Figure 4: Amplification with conserved and specific oligonucleotides. Lane 1, molecular weight marker. Lane 2 and 3 corresponds to amplification with MA1–MA2 conserved primers, using DNA samples from San Quintín and La Salina, respectively. Lane 4 and 5, corresponds to amplification with DBs-MA2 and DSs-MA2 specific-conserved oligonucleotides, using both β-carotene hyperproducers strains.

Mentions: Morphological identification demonstrated that red environmental samples contained Dunaliella strains (data not shown). Microscopic species differentiation was not possible due great visual similarities among Dunaliella species. Molecular characterization was carried out to obtain 18S rDNA fingerprinting profile from isolated strains. Standard PCR protocol using 18S rDNA gene conserved primers [5,11], was successfully utilized to amplify target rDNA region from the isolated Dunaliella strains. Two different sizes of DNA products were found in PCR reactions, using the same pair of conserved primers. Fragments around ~2500 bp long were found in samples coming from San Quintín B.C. In addition, fragments around ~2100 bp long were found in samples from La Salina (Fig. 4). In previous works, Wilcox and coworkers in 1992 demonstrated that microalgae strains with two introns inside 18S rDNA have a gene size of ~2500 bp long. Additionally, the same work showed that strains with one intron within the 18S rDNA have a gene size of ~2100 bp long. In this sense, PCR products amplified from red lagoons in Baja, Mexico, belong to D. salina/bardawil (~2500 bp) and "D. salina var Teod" (~2100 bp) respectively [5,11].


DNA fingerprinting differentiation between beta-carotene hyperproducer strains of Dunaliella from around the world.

Olmos J, Ochoa L, Paniagua-Michel J, Contreras R - Saline Syst. (2009)

Amplification with conserved and specific oligonucleotides. Lane 1, molecular weight marker. Lane 2 and 3 corresponds to amplification with MA1–MA2 conserved primers, using DNA samples from San Quintín and La Salina, respectively. Lane 4 and 5, corresponds to amplification with DBs-MA2 and DSs-MA2 specific-conserved oligonucleotides, using both β-carotene hyperproducers strains.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2710335&req=5

Figure 4: Amplification with conserved and specific oligonucleotides. Lane 1, molecular weight marker. Lane 2 and 3 corresponds to amplification with MA1–MA2 conserved primers, using DNA samples from San Quintín and La Salina, respectively. Lane 4 and 5, corresponds to amplification with DBs-MA2 and DSs-MA2 specific-conserved oligonucleotides, using both β-carotene hyperproducers strains.
Mentions: Morphological identification demonstrated that red environmental samples contained Dunaliella strains (data not shown). Microscopic species differentiation was not possible due great visual similarities among Dunaliella species. Molecular characterization was carried out to obtain 18S rDNA fingerprinting profile from isolated strains. Standard PCR protocol using 18S rDNA gene conserved primers [5,11], was successfully utilized to amplify target rDNA region from the isolated Dunaliella strains. Two different sizes of DNA products were found in PCR reactions, using the same pair of conserved primers. Fragments around ~2500 bp long were found in samples coming from San Quintín B.C. In addition, fragments around ~2100 bp long were found in samples from La Salina (Fig. 4). In previous works, Wilcox and coworkers in 1992 demonstrated that microalgae strains with two introns inside 18S rDNA have a gene size of ~2500 bp long. Additionally, the same work showed that strains with one intron within the 18S rDNA have a gene size of ~2100 bp long. In this sense, PCR products amplified from red lagoons in Baja, Mexico, belong to D. salina/bardawil (~2500 bp) and "D. salina var Teod" (~2100 bp) respectively [5,11].

Bottom Line: In this work, we applied our intron-sizing method to compare the 18S rDNA fingerprint between D. salina (CCAP 19/18), D. salina/bardawil (UTEX LB2538) and beta-carotene hyperproducing strains of Dunaliella isolated from salt saturated lagoons in Baja, Mexico.In Baja Mexico we found D. salina and D. salina/bardawil species by using intron-sizing-method.The National Center for Biotechnology Information (NCBI) Dunaliella 18S rDNA gene sequences were analyzed with our methodology and extraordinary correlation was found with experimental results.

View Article: PubMed Central - HTML - PubMed

Affiliation: Molecular Microbiology Laboratory, Centro de Investigación Científica y de Educación Superior de Ensenada (CICESE), Department of Marine Biotechnology, Ensenada, B.C, México.

ABSTRACT

Background: Dunaliella salina is the most important species of the genus for beta-carotene production. Several investigations have demonstrated that D. salina produces more than 10% dry weight of pigment and that the species grows in salt saturated lagoons. High plasticity in the green stage and the almost indistinguishable differences in the red phase make identification and differentiation of species and ecotypes very difficult and time consuming.

Results: In this work, we applied our intron-sizing method to compare the 18S rDNA fingerprint between D. salina (CCAP 19/18), D. salina/bardawil (UTEX LB2538) and beta-carotene hyperproducing strains of Dunaliella isolated from salt saturated lagoons in Baja, Mexico. All hyperproducer strains reached beta-carotene levels of about 10 pg/cell. Optical microscopy did not allow to differentiate between these Dunaliella strains; however, 18S rDNA fingerprinting methodology allowed us to differentiate D. salina from D. salina/bardawil.

Conclusion: In Baja Mexico we found D. salina and D. salina/bardawil species by using intron-sizing-method. The National Center for Biotechnology Information (NCBI) Dunaliella 18S rDNA gene sequences were analyzed with our methodology and extraordinary correlation was found with experimental results.

No MeSH data available.