Development and molecular characterization of genic molecular markers for grain protein and calcium content in finger millet (Eleusine coracana (L.) Gaertn.)

Finger millet (Eleusine coracana (L.) Gaertn), holds immense agricultural and economic importance for its high nutraceuticals quality. Finger millets seeds are rich source of calcium and its proteins are good source of essential amino acids. In the present study, we developed 36 EST-SSR primers for the opaque2 modifiers and 20 anchored-SSR primers for calcium transporters and calmodulin for analysis of the genetic diversity of 103 finger millet genotypes for grain protein and calcium contents. Out of the 36 opaque2 modifiers primers, 15 were found polymorphic and were used for the diversity analysis. The highest PIC value was observed with the primer FMO2E33 (0.26), while the lowest was observed FMO2E27 (0.023) with an average value of 0.17. The gene diversity was highest for the primer FMO2E33 (0.33), however it was lowest for FMO2E27 (0.024) at average value of 0.29. The percentage polymorphism shown by opaque2 modifiers primers was 68.23 %. The diversity analysis by calcium transporters and calmodulin based anchored SSR loci revealed that the highest PIC was observed with the primer FMCA8 (0.30) and the lowest was observed for FMCA5 (0.023) with an average value of 0.18. The highest gene diversity was observed for primer FMCA8 (0.37), while lowest for FMCA5 (0.024) at an average of 0.21. The opaque2 modifiers specific EST-SSRs could able to differentiate the finger millet genotypes into high, medium and low protein containing genotypes. However, calcium dependent candidate gene based EST-SSRs could broadly differentiate the genotypes based on the calcium content with a few exceptions. A significant negative correlation between calcium and protein content was observed. The present study resulted in identification of highly polymorphic primers (FMO2E30, FMO2E33, FMO2-18 and FMO2-14) based on the parameters such as percentage of polymorphism, PIC values, gene diversity and number of alleles.

[1]  P. Sofi,et al.  Review article: Quality protein maize (QPM): Genetic manipulation for the nutritional fortification of maize , 2009 .

[2]  W. Ooghe,et al.  Amino acid profiles after sprouting, autoclaving, and lactic acid fermentation of finger millet (Eleusine coracan) and kidney beans (Phaseolus vulgaris L.). , 2000, Journal of agricultural and food chemistry.

[3]  K. Sahrawat,et al.  Genetic diversity for grain nutrients contents in a core collection of finger millet (Eleusine coracana (L.) Gaertn.) germplasm , 2011, Field Crops Research.

[4]  J. Sambrook,et al.  Molecular Cloning: A Laboratory Manual , 2001 .

[5]  B. Leikind,et al.  An Assessment of , 2011 .

[6]  R. C. Babu,et al.  Assessment of Genetic Diversity among Finger Millet (Eleusine coracana (L.) Gaertn.) Accessions using Molecular Markers , 2007, Genetic Resources and Crop Evolution.

[7]  .. S.Chethan,et al.  Finger Millet Polyphenols: Characterization and their Nutraceutical Potential. , 2007 .

[8]  Masaru Tomita,et al.  A novel feature of microsatellites in plants: a distribution gradient along the direction of transcription , 2003, FEBS letters.

[9]  Kejun Liu,et al.  PowerMarker: an integrated analysis environment for genetic marker analysis , 2005, Bioinform..

[10]  J. Bennetzen,et al.  Assessment of genome origins and genetic diversity in the genus Eleusine with DNA markers. , 1995, Genome.

[11]  P. Shewry,et al.  Seed storage proteins: structures and biosynthesis. , 1995, The Plant cell.

[12]  P. Arruda,et al.  The Role of Opaque2 in the Control of Lysine-Degrading Activities in Developing Maize Endosperm , 1999, Plant Cell.

[13]  D. Labuda,et al.  Genome fingerprinting by simple sequence repeat (SSR)-anchored polymerase chain reaction amplification. , 1994, Genomics.

[14]  K. Livak,et al.  DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. , 1990, Nucleic acids research.

[15]  P. Vos,et al.  AFLP: a new technique for DNA fingerprinting. , 1995, Nucleic acids research.

[16]  M. B. Sørensen,et al.  Transcriptional and post-transcriptional regulation of gene expression in developing barley endosperm , 1989, Molecular and General Genetics MGG.

[17]  W. F. Thompson,et al.  Rapid isolation of high molecular weight plant DNA. , 1980, Nucleic acids research.

[18]  W. Martins,et al.  WebSat ‐ A web software for microsatellite marker development , 2009, Bioinformation.

[19]  F. Rohlf,et al.  NTSYS-pc Numerical Taxonomy and Multivariate Analysis System, version 2.1: Owner manual , 1992 .

[20]  Thomas Lübberstedt,et al.  Functional markers in plants. , 2003, Trends in plant science.

[21]  R. Saini,et al.  Efficiency of RAPD, SSR and Cytochrome P450 gene based markers in accessing genetic variability amongst finger millet (Eleusine coracana) accessions , 2010, Molecular Biology Reports.

[22]  Use of SSR, RAPD markers and protein profiles based analysis to differentiate Eleusine coracana genotypes differing in their protein content , 2012, Molecular Biology Reports.

[23]  W. Köhler,et al.  Population Genetics: Aspects of Biodiversity , 2003 .

[24]  C L L Gowda,et al.  Developing a Mini‐Core Collection in Finger Millet Using Multilocation Data , 2010 .

[25]  L. Parducci,et al.  Species identification in seven small millet species using polymerase chain reaction-restriction fragment length polymorphism of trnS-psbC gene region. , 2001, Genome.

[26]  A. Jacobson,et al.  Phylogenetic relationships in Alisma (Alismataceae) based on RAPDs, and sequence data from ITS and trnL , 2007, Plant Systematics and Evolution.

[27]  L. Parducci,et al.  Species identification in seven small millet species using polymerase chain reaction - Restriction fragment length polymorphism of trnS-psbC gene region , 2001 .

[28]  K. Hilu,et al.  Ribosomal DNA variation in finger millet and wild species of Eleusine (Poaceae) , 1992, Theoretical and Applied Genetics.

[29]  Liqing Zhang,et al.  Distinct patterns of SSR distribution in the Arabidopsis thaliana and rice genomes , 2006, Genome Biology.

[30]  M. Heun,et al.  Barley microsatellites: allele variation and mapping , 1995, Plant Molecular Biology.

[31]  Hari D. Upadhyaya,et al.  Development of Core Subset of Finger Millet Germplasm Using Geographical Origin and Data on 14 Quantitative Traits , 2006, Genetic Resources and Crop Evolution.

[32]  E. Mertz,et al.  Mutant Gene That Changes Protein Composition and Increases Lysine Content of Maize Endosperm , 1964, Science.

[33]  Richard D. Thompson,et al.  The transcriptional activatorOpaque-2 controls the expression of a cytosolic form of pyruvate orthophosphate dikinase-1 in maize endosperms , 1996, Molecular and General Genetics MGG.

[34]  David B. Goldstein,et al.  Microsatellites: Evolution and Applications , 1999 .

[35]  V. Yadav,et al.  Comparative evaluation of genetic diversity using RAPD, SSR and cytochrome P450 gene based markers with respect to calcium content in finger millet (Eleusine coracana L. Gaertn.) , 2010, Journal of Genetics.