Ekspresija gena za ribozom-inaktivirajući protein (SoRIP2) kao marker za analizu embriogenog potencijala spanaća (Spinacia oleracea L. cv. Matador) in vitro
Expression of gene encoding ribosome-inactivating protein (SoRIP2) as a marker for the analysis of spinach (Spinacia oleracea L. cv. Matador) embryogenic potential in vitro
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Somatic embryogenesis was induced from apical sections of the lateral roots ofspinach seedlings (Spinacia oleracea L. cv. Matador), cultivated in vitro on Murashige &Skoog induction medium supplemented with 20μM α-naphthaleneacetic acid (NAA) and 5μM gibberellic acid (GA3). The capacity for somatic embryo formation was tested in 30randomly chosen seedlings (lines). Results indicated that the regeneration potential is veryvariable among the seedlings, with the frequency of regeneration ranging from 0.3% to100% and the mean somatic embryo (SE) number per explant ranging from 0.001 to 9.96.Only 4 out of 30 lines displayed high regeneration frequency (85-100%), with the totalnumber of 227-347 SEs regenerated from 30 root sections over a 12-week period. Only oneline regenerated during the first subculture (4 weeks). Results indicate that the lines withhigh embryogenic potential are very rare within the population of cultivar "Matador".SEs isolated from a single seedling were maintained as a line on medium with 5μMkinetin. Apical shoots, isolated from the same seedlings, which have been used for theinduction of regeneration, were cultivated separately in test tubes, on plant growthregulator-free medium. Plants derived from seedlings’ apical shoots and SEs flowered,self-fertilised and set seeds in vitro. Thus, four generations of descendent seedlings,designated as S1-S4, were obtained by self-fertilisation and germination of the collectedseeds in vitro. Roots of these seedlings were used for the induction of SE regeneration andtheir embryogenic capacities were compared to parental lines. The embryogenic capacitiesof S1 and S2 lines were also very variable. By selection of offspring with high embryogenicresponse and through further cycles of self-fertilisation, three S3 and one S4 line withextremely high embryogenic capacities, 33-35 fold higher than in the mother-plant, wereobtained. The regeneration frequencies of these lines were approximately 100%, and1339-2181 SEs were isolated from 30 root sections per seedling over a 12-week period. Alllines regenerated during the first subculture, and 2-4 weeks earlier compared to parentallines. Root sections of plants regenerated from SE (regenerants) exhibited similarregeneration capacity as the respective seedling from which they originated...
Keywords:Spinacia oleracea; Genotype; Somatic embryogenesis; Photoperiod; Lightintensity; Gibberellins; SoRIP2; SoGA20-ox1; SoGA3-ox1; SoGA2-ox1; Molecular markers; Gene expression
Source:University of Belgrade, Faculty of Biology, 2015, 1-185