Molecular analysis of plants regenerated from embryogenic cultures of hybrid sugarcane cultivars (Saccharum spp.)

Abstract

The genomic stability of tissue culture regenerants of sugarcane (Saccharum spp. hybrids, cvs 'CP721210', 'CP68-1067' and 'B43-62') was analyzed by DNA restriction fragment length polymorphism (RFLP). Plants regenerated from calli, cell suspensions, cryopreserved cell suspensions and protoplasts were used. Total DNA isolated from 19 different sources was digested with EcoRI, HindIII, BamHI, BamHI, EcoRI and PstI and probed with six known maize mitochondrial genes (coxI, coxII, atpA, atp6, atp9 and rrn18-rrn5), three random maize mitochondrial cosmid clones, two random maize chloroplast cosmid clones and a wheat Nor locus clone. Hybridization patterns indicated that the variation observed was minor and appeared only in the secondcycle regenerants. No differences were observed among the three cultivars and the regenerants from calli, suspension culture, cryopreserved suspension culture and protoplasts. Mitochondrial DNA (mtDNA) isolated from 'CP72-1210' plants and its embryogenic cell suspensions, and bulk samples from all 'CP72-1210' regenerants pooled together were digested with EcoRI, HindIII, PstI, BamHI and SalI and probed with three recombinationally active wheat mtDNA clones, K', K3 and X2. No variation in the mtDNA restriction patterns was observed between the 'CP72-1210' plants and its regenerants. However, restriction pattern variation was observed only from EcoRI digestion, and hybridization patterns of K3, K' and X2 revealed minor variations in the mtDNA of cell suspensions when compared with the DNA of the 'CP72-1210' plant. Except for a qualitative variation detected by the X2 probe and minor stoichiometric variations detected by the K3 probe, sugarcane DNAs were found to be stable after plant regeneration.

Micropropagation of Ocimum kilimandscharicum Guerke (Labiatae)

“…A major problem encountered in cells grown in vitro is genetic variation. In many plant species, PCR-based RAPD markers have been used to confirm the clonal fidelity and genetic stability of plants grown in tissue culture and donors (Chawdhury and Vasil, 1993;Rani et al, 1995;Rout et al, 1998;Das and Pal, 2005;Dewir et al, 2005;Chaudhuri et al, 2008;Bhattacharya et al, 2009). We made the DNA fingerprinting profiles of Ocimum kilimandscharicum plants regenerated in culture from nodal stem segment explants and the respective donor plants, using 10 RAPD primers.…”

Untitled

“…However, some bands were amplified from the F1 hybrid but not from parents. The reason could be the complex heredity background of each plant heterosis [28][29][30] . Additionally, this residual heterozygosity could account for the occasional occurrence that some true hybrids exhibited the absence of the female or www.scienceasia.org male specific-parent markers with RAPD primers.…”

Cryopreservation of Tropical Plant Germplasm with Vegetative Propagation - Review of Sugarcane (Saccharum spp.) and Pineapple (Ananas comusus (L.) Merrill) Cases

“…By contrast, only limited information is available concerning the stability of plants regenerated from cryopreserved material. RFLP analysis did not reveal any difference that could be attributed to cryopreservation between plants of one sugarcane variety produced from control and cryopreserved calluses (Eksomtramage et al, 1992) or cell suspensions (Chowdhury & Vasil 1993). Plants produced from control and cryopreserved shoot tips of one variety were similar as regards pattern of two isoenzymatic systems (Paulet et al, 1993) and six agronomic traits observed during their early growth in vivo (Gonzalez-Arnao et al, 1999).…”