Rice Anther Culture Research Articles

Phenylacetic acid stimulation of direct shoot formation in anther and somatic tissue cultures of rice (Oryza saliva L.)

AbstractThe effect of phenylacetic acid (PAA) on rice (Oryza saliva L.) anther culture was investigated with six genotypes, using 2,4‐D as control. In the two‐step culture protocol, replacing 2, 4‐D with PAA in the induction medium did not influence callus induction but significantly improved the shoot differentiation from callus, particularly in the indica cultivar Teqing. The anther‐derived calli of all genotypes regenerated shoots directly on the callus induction medium containing PAA. Most of the directly‐regenerated plantlets had well‐developed root systems and were therefore readily transplanted into soil. The improved shoot differentiation potential and the frequency of direct regeneration depended on genotype, basal medium and PAA concentration. The one‐step green shoot regeneration frequencies obtained were 1.98% with the indica cultivar ‘129’, 1.5% with the indica × japonica hybrid ‘Teqing/02428’ (F1), and 1.98% with the indica × indica hybrid ‘Waiyin 2/C.B.’ (F1). The PAA‐based one‐step method was most effective on the anther culture of indica genotypes. Three DH populations have been constructed from hybrids (F1) via one‐step culture. PAA also enhanced the one‐step plantlet formation in rice somatic tissue culture.

Genetic studies of anther culture ability in rice (Oryza sativa)

Inheritance of three anther and culture characters, callus induction, green plant regeneration and culture efficiency was studied using incomplete diallel crosses with a gamete model. It was suggested that callus induction was mainly controlled by gametic additive effects and with less effect of the maternal effects. Green plant regeneration was mainly determined by maternal effects with less influence of gametic additive effects. Culture efficiency was controlled by gametic additive, maternal and cytoplasmic effects. Cultivar Lunhui 422 showed positive genetic effects for all three traits and was a very good parent for rice anther culture breeding. Significant positive heterosis was observed for callus induction. Both gametic additive and maternal correlations contributed to the significant genotypic and phenotypic correlations between callus induction and green plant regeneration suggesting these two traits to be linked.

Transcription of tRNA Genes from a Large-Scale Plastid DNA Deletion Clearly Reveals the Action of Nuclear-Encoded RNA Polymerase in the Plastid

Summary We have recently characterized large-scale deleted plastid DNAs in calli derived from rice (Oryza = sativa) anther culture. The size and location of these deletions differed between the calli. In this study the conformation of the largest plastid DNA deletion of callus A 10 was studied using neutral-alkaline, two-dimensional, agarose-gel electrophoresis. This clearly revealed that its DNA had the same conformation as that of the previously described callus A5: linear molecules with a hairpin structure at both termini and existing in both monomer and multimer forms linked to form head-to-head and tail-to-tail concatemers. The region retained by both A10 and A5 was only about 3kbp in size and encoded trnfM , trnG , ϕ trnI , trnI , trnT , and trnE Transcripts from these deleted ptDNAs were identified by RNA gel blotting using oligonucleotide DNA probes. These results clearly indicated the involvement of nuclear-encoded RNA polymerase in the transcription of the tRNA genes because these plastid genomes lack nearly all the genes encoding their own translational apparatus (i.e. rRNA and tRNA). The A10 plastid also lacks all RNA polymerase genes ( rpoA, B and C ). The function of the large-scale plastid DNA deletion is discussed.

Androgenesis of highly recalcitrant rice genotypes with maltose and silver nitrate

Callus formation per anther of highly recalcitrant rice genotypes increased from 6.3 to 20.6% when sucrose was replaced by maltose and AgNO 3 was added to the induction medium ( P = 0.01). The formation of green plants increased by 15 times when maltose in the induction medium was raised from 29 to 351 mM. On the other hand, glucose drastically reduced callus induction, and combinations of maltose and mannitol were less inductive than equiosmolal concentrations of maltose. No differences were noted when sucrose or maltose and silver nitrate were included in the regeneration medium, indicating that in rice anther culture the type of sugar and the anti-ethylene compound mainly affects callus induction and embryogenesis, but not plant development. These factors were investigated on diverse rice genotypes that represented a range of genetic diversity used in breeding.

Efficiency of Different Media Used in Anther Culture of Hybrid Rice

Efficiency of Different Media Used in Anther Culture of Hybrid Rice

Rice anther culture: callus initiation and androclonal variation in progenies of regenerated plants

Anthers from rice (Oryza sativa L.) subspecies japonica initiated more callus than their indica or indica x japonica counterparts. A mild stress, either by slow desiccation or heat shock, prior to the plating of anthers enhanced the ability to initiate callus. Slow dessication of anthers enhanced the ability of the japonica anthers to initiate callus even in medium that was supplemented with NaCl. The ability to initiate callus by the anthers plated on NaCl-supplemented medium decreased as the NaCl level in the medium increased. Among the regenerated plants 2.5% were albino and another 2% were haploid. Androclonal variation for tiller numbers, shoot height, plant dry matter and flowering were noticed in the progenies of the regenerated plants.

Observations and Suggestions for Improving Somatic Hybridization by Plant Protoplast Isolation, Fusion, and Culture

De (Triticum aestivum): In vitro production and utilization of doubled haploids, p. 101–124. In: Y.P.S. Bajaj (ed.). Biotechnology in agriculture and forestry. vol. 12: Haploids in crop improvement I. Springer-Verlag, Berlin. Powell, W., P.D.S. Caligari, and J.M. Dunwell. 1986. Field performance of lines derived from haploid and diploid tissues of Hordeum vulgare. Theor. Applied Genet. 72:458–465. Reed, S.M. and E.A. Wernsman. 1989. DNA amplification among antherderived doubled haploid lines of tobacco and its relationship to agronomic performance. Crop Sci. 29:1072–1076. 401 Research Group, Laboratory of Plant Cell and Tissue Culture, Institute of Genetics, Academia Sinica. 1975. Primary study on induction of pollen plants of Zea mays. Acta Genet. Sin. 2:143. Research Laboratory of Breeding. 1981. A preliminary study on the heredity and vitality of the progenies of tobacco pollen plants, p. 223–225. In: H. Hul (ed.). Plant tissue culture. Proc. Symp. Plant Tissue Culture, Peking, 1978. Pitman Publishing, London. Rufty, R.C., E.A. Wernsman, C.E. Main, and G.V. Gooding, Jr. 1990. Registration of NC-BMR 42 and NC-BMR 90 germplasm lines of tobacco. Crop Sci. 30:241–242. Schaeffer, G.W., F.T. Sharpe, Jr., and P.B. Cregan. 1984. Variation for improved protein and yield from rice anther culture. Theor. Applied Genet. 67:383–389. Simmonds, N.W. 1979. Principles of crop improvement. Longman, London. Thomas, E., F. Hoffman, and G. Wenzel. 1975. Haploid plantlets from microspores of rye. Z. Pflanzenzuchtg. 75:106–113. Wang, Y., C. Sun, C. Wang, and N. Chien. 1973. The induction of the pollen plantlets of Triticale and Capsicum annuum from anther culture. Sci. Sinica 16:147–151. Wenzel, G., F. Hoffmann, and E. Thomas. 1977. Anther culture as a breeding tool in rape. I. Ploidy level and phenotype of androgenetic plants. Z. Pflanzenzuchtg. 78:149–155. Wenzel, G., O. Schieder, T. Przewozny, S.K. Sopory, and G. Melchers. 1979. Comparison of single cell culture derived Solanum tuberosum L. plants and a model for their application in breeding programs. Theor. Applied Genet. 55:49–55. Winzeler, H., J. Schmid, and P.M. Fried. 1987. Field performance of androgenetic, doubled haploid spring wheat lines in comparison with lines selected by the pedigree system. Plant Breeding 99:41–48. Wu, J. 1986. Breeding haploid corn by anther culture, p. 149–161. In: H. Hu and H. Yang (eds.). Haploids of higher plants in vitro. Springer-Verlag, Berlin. Yang, X. and H. Fu. 1989. Hua-03—A high protein indica rice. Intl. Rice Res. Nwsl. 14(3):14–15. Zhu, D. and X. Pan. 1990. Rice (Oryza sativa L.): Guan 18—An improved variety through anther culture, p. 204–211. In: Y.P.S. Bajaj (ed.). Biotechnology in agriculture and forestry 2: Haploids in crop improvement I. Springer-Verlag, Berlin. Zhao, Y., X. He, J. Wang, and W. Liu. 1990. Anther culture 28—A new diseaseresistant and high-yielding variety of winter wheat, p. 353–362. In: Y.P.S. Bajaj (ed.). Biotechnology in agriculture and forestry 13: Wheat. SpringerVerlag, Berlin.

Evaluation of Rice Anther Culture Lines for Tolerance to the Rice Water Weevil (Coleoptera: Curculionidae)

Preliminary field evaluation of 43 rice anther culture lines in 1990 for resistance to the rice water weevil, Lissorhoptrus oryzophilus Kuschel, identified several lines with potential for tolerance. The seven most promising lines (952836, 953508, 953509, 953510, 953511, 953527, and 953541) were evaluated for tolerance in 1991 and 1992. The parameters used to assess tolerance included rice water weevil larval population, root pruning damage, plant height, and grain yield. Significant differences were found among the lines tested with regard to rice water weevil larval populations, root damage ratings, plant height, and grain yields. The comparison of insecticide treated and untreated plots indicated that lines 952836 and 953527 exhibited moderate levels of tolerance to the rice water weevil. Tolerance in the lines was expressed, in part, as the ability to recover from root pruning damage and to produce high grain yields.

Identification of Ploidy Level of the Regenerated Plants by Anther Culture in Rice.

Identification of Ploidy Level of the Regenerated Plants by Anther Culture in Rice.

Utilisation of anther culture in rice (Oryza sativa L.) breeding in Italy

Increasing market demand has led to efforts to select for non-sticky, long-grain rice varieties suited to the Italian environment. The variability of recombinant populations derived from crosses between Italian and USA genotypes was exploited. Together with the conventional breeding methods, in vitro anther culture was used to obtain homozygous lines from crosses more quickly. Moreover, hybrid lines were obtained from crosses otherwise impossible to exploit owing to sterility in the F, generation (namely those resulting from crosses involving semi-dwarf USA varieties). Plant regeneration was obtained from 6 F, hybrids whose parents were chosen for their grain quality or their suitability to the Italian environment. About 5500 plantlets were regenerated. Of these, 1000 were albino, and about 50% of the green ones died after transplanting into soil; 65% of the surviving plants were sterile, so about 1000 progeny were obtained (30-445 for each cross). Progeny with desirable characteristics were evaluated for agronomic traits. On the basis of these results, limits and possibilities of anther-culture breeding in rice are discussed.

Heterozygous diploid plants regenerated from anther culture of F1 rice plants

Plants derived from anther culture are theoretically haploid, but diploid plants are also known to arise. Anther culture-derived diploid plants are usually homozygous and are believed to be due to spontaneous doubling of chromosomes in either microsporocytes or callus cells during the culture process. However, heterozygous diploid regenerants may also arise from a) regeneration from cultured somatic cells, b) mutation occurring during or after a spontaneous doubling event, c) fusion of unlike haploid cells in chimeric callus, and d) regeneration from diploid microsporocytes resulting from aberrant meioses. This study was designed to elucidate the frequency and origin of diploid regenerants from rice anther culture. Regenerants were obtained from 11 F1 genotypes. Progeny testing detected heterozygosity in 7 out of 211 regenerants. Each of the heterozygous regenerants were from ‘Calrose 76’/waxy ‘M-101’, Half of the diploid regenerants from this cross were heterozygous. No heterozygous regenerants arose from the other 10 F1 genotypes. Progeny testing indicated that two of the heterozygous regenerants were as heterozygous as the F1 plants for three parental characters. The other five regenerants exhibited decreased levels of heterozygosity. One of the heterozygous regenerants exhibited evidence of mutation for a non-parental character. However, mutation is an unlikely cause of the observed high levels of parental-type heterozygosity. No evidence for the occurrence of chimeric callus was detected, making this an unlikely cause as well. The most likely origin of the observed partial heterozygosity is regeneration from diploid microspores, which could also produce plants exhibiting complete parental-type heterozygosity.

Genetic studies of rice (Oryza sativa L.) anther culture response

Anthers of two rice (Oryza sativa L.) varieties, Lunhui 422 (P1) and Jinzao 5 (P2), their F1, F2 and first backcross generation-F1 x Lunhui 422 (B1), and F1 x Jinzao 5 (B2)-were cultured in L8 medium to study the inheritance of rice anther culturability using generation mean analysis. Significant effects of generation were observed for the four traits measured: anther response (%), frequency of callus induction (%), frequency of callus differentiation (%) and culture efficiency (%). Variation among the generations was similar for all traits: significant differences were found among six generations and the means of the P2 and B2 were significantly lower than those of the other generations. The frequency of callus differentiation showed a nonsignificant difference among the P1, F1, F2 and B1 generations which had slightly high values than the P2 and B2. Additive genetic variance (VA) was higher than non-additive genetic variance (VD) for anther response and frequency of callus induction. However, AV was lower than VD for frequence of callus differentiation and culture efficiency, VA was significant for all traits except for the culture efficiency, and VD was nonsignificant for all traits except for the frequency of callus differentiation. On the other hand, environmental variation (VE) was significant for the 4 traits. Narrow-sense heritability estimates were 95.52%, 82.19% and 13.54% for anther response, frequency of callus induction and culture efficiency, respectively.

Genetic Variation of Characters induced by Anther Culture and Seed Cullus Culture of Rice

Genetic Variation of Characters induced by Anther Culture and Seed Cullus Culture of Rice

Genealogical Study on Callus Formation Ability in Anther Culture of Rice Variety, Koshihikari.

Callus formation ability in anther culture of rice (Oryza sativa L.) variety, Koshihikari, was investigated genealogically. First, to ascertain the varietal difference in the quality of callus in anther culture of rice, 31 main varieties in Oita Prefecture were studied. There was a significant varietal differ-ence, and they were divided into good callus formation varieties and poor callus formation varieties. The former formed pale yellow and vigorously growing calli. The latter formed brown and poorly growing calli. These results showed that the quality of callus in anther culture of rice could be characterized by the color and growth of callus.

Interaction of Media for Callus Induction and for Plant Regeneration in Rice Anther Culture

Interaction of Media for Callus Induction and for Plant Regeneration in Rice Anther Culture

Genetic Analysis of Bacterial Blight (BB) Resistance in Rice Anther Culture Progenies

Genetic Analysis of Bacterial Blight (BB) Resistance in Rice Anther Culture Progenies

Efficient Induction of Diploidized Plants in Anther Culture of Rice by Colchicine Pretreatment of Cold-Preserved Spikes.

Effects of colchicine pretreatment of cold-preserved spikes on the increase in ratio of diploidized fertile regenerants in anther culture of rice were studied. After 10 days of storage at 10°C, cut ends of the spikes including uninucleate pollen grains were dipped into colchicine solution at the same temperature. When the spikes were treated with 0.8% colchicine solution for 3 days, more than 50% of the regenerants, obtained in subsequent anther culture, were fertile.

Diallel analysis of callus formation ability in anther culture of rice.

イネの葯培養におけるカルス形成率の遺伝様式を調べるために,形成率の異なる日本型,インド型の水稲ならびに陸稲の計6品種の総当たり交雑を行い,雑種F1の葯のカルス形成率のダイアレル分析を行った.カルス形成率は,水・陸稲とも日本型水稲品種が高く,インド型品種がかなり低かった。カルス形成率の遺伝的変異は、主として遺伝子の相加的効果によることが推測された.その平均優性度(√H1/D)はO.39であった.カルス形成率の高いほうが劣性であった.カルスをほとんど形成しない I Kong Pao は完全優性親カルス形成率の高い北陸100号は完全劣性親と推察された.その他の現品種については優性遺伝子と劣性遺伝子を併有していると考えられた.分散分析の結果,親聞及び特定組合せの優性効果に有意差が認められた.優性効果発現には,カルス形成率の低いインド型陸稲品種赤米dがとくに関与しており,この品種を除いた副ダイアレル表の分析では,親間および特定組合せ間優性効果が有意でなくなり,平均優性度も0.19と減少した.正逆交雑に差は認められず,細胞質または母体効果はカルス形成率に影響していないと考えられた.

Diallel Analysis of Callus Induction and Green‐Plant Regeneration in Rice Anther Culture

Low efficiencies of callus induction and green‐plant regeneration have limited the application of anther culture in breeding programs. This study was conducted to determine the inheritance of callus induction and plant regeneration in rice (Oryza sativa L.) anther culture. Sixteen genotypes derived from a four‐parent diallel mating set of two Japonica cultivars (Taipei 309 and Taipei 177) and two indica cultivars (IR36 and Basmati 370) were evaluated for their callus induction and plant regeneration abilities. For callus induction, the effects of genotype and genotype ✕ medium interaction were significant (P < 0.05). Similarly, genotype and abscisic acid (ABA) level were found to be significant (P < 0.05) for green‐plant production. The japonica cultivars generally had a higher percent callus induction and plant regeneration, and number of calli producing green plants than did the indica cultivars. Similar modes of inheritance were observed for both characters. Combining ability analysis demonstrated the predominance of additive gene effects in the control of both characters with the japonica cultivars having higher combining ability for green‐plant regeneration. Reciprocal effects were not significant (P > 0.05), suggesting the absence of maternal control on the characters. Further analysis showed gene action to be partially dominant with high response being highly recessive and controlled by a few genes. There were no indications of inter‐allelic interaction.

Varietal differences in the response to low temperature treatment for callus formation in anther culture of rice.

Callus-forming ability in anther culture was studied in 7 Japanese rice cultivars, consisting of chilling-resistant and susceptible ones at the pollen-forming stage, after treatment of the panicles at 5, 10 and 15°C for 8, 16, 24, 32 and 40 days. The rate of callus formation (percentage of the number of callus-formlng anthers to that of anthers plated) differed depending on the varieties as well as the temperature and duration of the treatment. The Hayayuki and Norin 20 cultivars in the Hokkaido area, formed a callus at a high rate by the 15°C treatment, while Somewake showed a good response to 10 and 5°C and to prolonged treatment. Hatsunishiki, Sasanishiki, Koshihikari and Norin 21 formed a callus at a fairly high rate by the 10 and 5°C treatments but at a low rate by the 15°C treatment. No significant correlation was observed between the callus formation rate and the chilling resistance in the cultivars. Varietal differences in the callus-forming response to temperature and duration of the pretreatment in anther culture were demonstrated.