In vitro selection and field responses of somaclonal variant plants of rice cv PR113 for drought tolerance

Drought is the major environmental stress that limits rice productivity worldwide. In vitro somaclonal variation using different selection agents has been used for crop improvement. Here, rice plants of cv PR113 were selected in vitro on 30, 50 and 70 g L^-1 polyethylene glycol 6,000 (PEG). Callus growth, proliferation, calli volume (first and second culture) and plantlet regeneration (third culture) were found to be decreased upto a certain level to acquire tolerance to PEG-induced drought. From the field data, 30 g L^-1 PEG lines showed higher vegetative growth (plant height, tiller number, leaf number, shoot weight and root growth) as compared with 50 g L^-1 PEG selected somaclone lines under limited irrigation. The yield parameters-panicle length, panicle weight, grains per panicle, 1,000-grain weight, grain yield per plant, harvest index and grain straw ratio were also higher in 30 g L^-1 PEG lines as compared with 50 g L^-1 PEG lines. The results, therefore indicate that 30 g L^-1 PEG selected somaclone lines were more suited than 50 g L^-1 PEG selected somaclone lines under stress as compared with WT. The finding suggests that rice cv PR113 somaclones generated on PEG are found to be drought tolerant under field condition with better yield.     

Menthol tolerant clones of Mentha arvensis: approach for in vitro selection of menthol rich genotypes

In vitro raised shoots of Mentha arvensis L. were screened for menthol tolerance level by growing them in media containing 0–100 g ml^–1 menthol. A total of 2850 regenerated shoots were step wise screened for menthol tolerance at the concentrations of 50 g ml^–1 followed by 60 and 70 g ml^–1. In this screening, only 30 individual regenerated shoots were able to survive. The clones from the primary screen were inoculated into rooting medium and, after rooting, transferred to pots in the greenhouse. Ultimately, these 30 menthol tolerant clones were multiplied and grown in the field in replicated plots of 2.5×2.5 m sizes. Twigs of 30 clones from the replicated trials were rechecked for tolerant phenotypes at a concentration of 70 g ml^–1 menthol wherein, these survived even after 7 days (secondary screening). These clones were checked for oil and menthol content and were found to be better than the control plants. Out of these 30 plants, five tolerated 80 g ml^–1 menthol (tertiary level screening) and were found to contain the highest amount of menthol per g leaf biomass. Molecular analysis through RAPD showed distinct variation in the profiles of these five plants, in comparison to the control. Using this method the relationship between the primer OPT 04, menthol tolerance and high menthol content character of the genotype was established. Further, a cultivar `Saksham' was released from the selections by CIMAP for superior performance.     

Selection in vitro for UV-tolerant sugar beet (Beta vulgaris) somaclones

With a reduced stratospheric ozone concentration, the generation of UV-tolerant plants may be of particular importance. Among different crop plants there is large variation in sensitivity to UV-B radiation. This study was undertaken to investigate the possibilities of using somaclonal variation and selection in vitro for improving UV-B tolerance in sugar beet (Beta vulgaris L.). Sugar beet callus was exposed to UV radiation (280–320 nm, 0.863–5.28 kJ m^-2 day^-1, unweighted) and resultant shoots were selected from surviving cells. After establishment of the plants, they were grown under either visible radiation (114 μmol m^-2 s^-1 PAR) or with the addition of UV radiation (6.3 kJ m^-2 day^-1 biologically effective UV-B). Screening of regenerants in vivo for tolerance to UV radiation was undertaken 10 months after termination of the UV selection pressure. Screening was done visually and by using a number of physiological parameters, including chlorophyll fluorescence induction, ultraweak luminescence, pigment analysis and total content of UV-screening pigments. A clear difference between the unselected and the UV-selected somaclones was observed when visually studying the UV damage and other leaf injury. The observations were supported by the ultraweak luminescence measurements. Unselected plants showed significantly greater damage when subjected to subsequent UV radiation as compared to the selected plants. The clones subjected to UV selection pressure displayed a significantly higher concentration of UV-screening pigments under subsequent UV radiation. The unselected plants under subsequent UV treatment showed a lower carotenoid concentration when compared to selected plants. However, no significant difference between treatments was found for chlorophyll a/b, or F/F_max, a measure of photosynthetic quantum yield.     

The influence of plant growth regulator concentrations and callus age on somaclonal variation in callus culture regenerants of strawberry

The effect of plant growth regulator concentrations and ageing of callus on the extent and nature of variation among callus culture regenerants of strawberry (Fragaria × ananassa) cv. Redcoat was examined. Plants regenerated from callus culture had reduced plant vigour, shorter petiole length and smaller leaf size, but more leaves and runners under greenhouse conditions. These responses appeared to be due to a physiological influence of plant growth regulators. No distinct phenotypic variants were observed at plant growth regulator concentrations in the range of 1–10 M each of BA and 2,4-d combination, but the highest concentration (20 M each) of this combination produced a high frequency (10%) of dwarf type variants. The dwarf nature of these variants was maintained in the runner plants produced by the primary regenerants. The plants regenerated from 8-week-old calli did not show any distinct morphological variants. However, a significant proportion of deformed leaf shape (6–13%) and yellow leaf (21–29%) variants was obtained among plants regenerated from 16 and 24-week-old calli. The primary regenerants of the leaf shape variants were established as chimeras. The chimeric plants produced runner progeny with normal plants and plants with completely distorted leaf morphology. Both leaf shape and yellow leaf variants remained stable through runner propagation. Isozyme analysis failed to distinguish any of the variants from the standard runner plants. Flow cytometric analysis indicated the aneuploid nature of leaf shape variants but it could not distinguish dwarf and yellow leaf variants from standard runner plants.     

A comparison of somatic chromosomal instability in tissue culture regenerants from Medicago media Pers

Two cultivars (Heinrichs, Reaver) and two breeding lines (Br1, Le1) from Medicago media were cultured in a media protocol consisting of a high concentration 2,4-D induction step. Regenerants were produced from all four stocks. Representative samples from each regenerant population along with the corresponding control population were cytologically analyzed for chromosomal and pollen abnormalities. While numerical changes in chromosome numbers were found in all regenerant populations, there was considerable variation between the four stock groups. Heteroploidy was observed for both hypo and hyper aneuploid regenerants, but there were no differences in pollen stainability between hypo and hyper aneuploid regenerants and euploid regenerants. Tissue culture regenerants generally produced a lower pollen stainability percent as compared to control populations grown from seeds. Gross and cryptic changes in chromosomes, or hormonal carry over effects or both were considered causes for poor pollen stainability in tissue culture regenerants. Cytological analyses indicate that the cultivar might play an important role in the cytological stability or instability of regenerant populations. Exploitation of this naturally existing situation to produce euploid regenerants for field experiments and to obtain gross cytological stability in somaclones is discussed.     

Plant regeneration from in vitro cultures of anthers and mature seeds of rice (Oryza sativa L.) cv. Basmati-370

Pollen plants were obtained from anther-derived calli of the indica rice variety Basmati-370. Anther-response (anthers producing pollen derived calli) and plant regeneration frequency from the pollen derived calli. was very low. Donor plants which flowered at the average max/min. temperature of 34.2°/23.3°C gave a significantly higher anther-response to in vitro techniques, than did those which flowered at 29.1°/16.4°C. Somatic callus induction and subsequent plant regeneration was readily obtained from mature seed embryos. While 2,4-D or 2,4,5-T (1 or 2 mg/l) proved highly efficient for callus induction, tryptophan (50 or 100 mg/l) induced a high frequency of green plants from the calli.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - 2,4,5-T 2,4,5-trichlorophenoxyacetic acid - NAA naphthaleneacetic acid - IAA indoleacetic acid - K kinetin - BA benzyladenine - Trp tryptophan - CW coconut water     

二棱大麦体细胞胚性无性系的建立

对１２个品种的二棱大麦（Ｈｏｒｄｅｕｍｄｉｓｔｉｃｈｕｍ２ｎ＝２４）幼穗和６个品种的幼胚在附加５００ｍｇ／Ｌ酪蛋白,１５０ｍｇ／Ｌ天门冬素和２ｍｇ／Ｌ２,４－Ｄ的ＭＳ培养基（Ｍ＿（３２０））中进行了离体培养,并诱导产生出了愈伤组织。把由Ｄ＿１幼胚所获得的愈伤组织转移到含２,４－Ｄ０．５ｍｇ／Ｌ（Ｍ＿（３０５））培养基中后诱导产生出了体细胞胚性愈伤组织,幼苗和根。筛选出了体细胞胚性无性系。在长达二年的继代培养中,该细胞系仍保持旺盛的再生和分化成苗能力。     

Selection of brown spot-resistant rice plants from Helminthosporium oryzae toxin-resistant calluses

Helminthosporium oryzae toxin induced electrolyte leakage from rice callus tissues and caused their browning and death. A virulent isolate of the pathogen invaded and colonised callus tissues rapidly, while a less virulent and a nonpathogenic isolate colonised calluses only weakly if at all. Addition of the toxin to calluses permitted colonisation of calluses by the nonpathogenic isolate. Four toxin-resistant calluses were selected and plants regenerated from two of these resistant calluses showed resistance to the pathogen. This resistance was heritable and stability of resistance was observed in the R₁, R₂ and R₃ generations.