Optimization of RT-PCR reactions in studies with genes of lignin biosynthetic route in Saccharum spontaneum



Document title: Optimization of RT-PCR reactions in studies with genes of lignin biosynthetic route in Saccharum spontaneum
Journal: Anais da Academia Brasileira de Ciencias
Database: PERIÓDICA
System number: 000420630
ISSN: 0001-3765
Authors: 1
1
1
Institutions: 1Universidade Estadual de Campinas, Instituto de Biologia, Campinas, Sao Paulo. Brasil
Year:
Season: Mar
Volumen: 90
Number: 1
Pages: 509-520
Country: Brasil
Language: Inglés
Document type: Artículo
Approach: Analítico, descriptivo
English abstract Saccharum spontaneum has been used for the development of energy cane a crop aimed to be used for the production of second-generation ethanol, or lignocellulosic ethanol. Lignin is a main challenge in the conversion of cell wall sugars into ethanol. In our studies to isolate the genes the lignin biosynthesis in S. spontaneum we have had great difficulty in RT-PCR reactions. Thus, we evaluated the effectiveness of different additives in the amplification of these genes. While COMT and CCoAOMT genes did not need any additives for other genes there was no amplification (HCT, F5H, 4CL and CCR) or the yield was very low (CAD and C4H). The application of supplementary cDNA was enough to overcome the non-specificity and low yield for C4H and C3H, while the addition of 0.04% BSA + 2% formamide was effective to amplify 4CL, CCR, F5H and CCR. HCT was amplified only by addition of 0.04% BSA + 2% formamide + 0.1 M trehalose and amplification of PAL was possible with addition of 2% of DMSO. Besides optimization of expression assays, the results show that additives can act independently or synergistically
Disciplines: Agrociencias
Keyword: Gramíneas,
Genética,
Caña de azúcar,
Mejoramiento genético,
Lignina,
Vías biosintéticas,
Regulación génica
Keyword: Gramineae,
Genetics,
Sugar cane,
Genetic improvement,
Lignin,
Biosynthetic ways,
Gene regulation
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