Transmission of a Bacillus thuringiensis cry3Aa transgene from diploid to tetraploid potato using 4x-2x hybridization: effect of ploidy increase on transgene expression and implications for TPS hybrid production

The study examined the effect of ploidy elevation through unreduced gametes on transgene expression in potato. Tetraploid transgenic progenies were obtained from one tetraploid potato cultivar crossed with 2n pollen producing diploid clones harbouring an exogenous transgene (cry3Aa). Both single‐ and multiple‐insert diploid transgenic lines that were regenerated by Agrobacterium tumefaciens leaf disc inoculation were used in crosses. A DAS‐ELISA system and no‐choice feeding bioassay enabled characterization of the parental lines as either ‘high’ or low’ expressers of the Cry3Aa protein. High Cry3Aa expression was observed for both single‐insert transgenic diploids and their 4x‐2x progeny. On the contrary, 68% of 4x‐2x progeny derived from a multiple‐insert, diploid transgenic had significantly reduced Cry3Aa expression compared with the parent, with 32% demonstrating nearly complete silencing of the transgene. Multiple copies of a transgene, like homologous native genes, may be susceptible to transgene silencing following polyploidization. Therefore, incorporation of exogenous transgenes into a true potato seed (TPS) production system is feasible if a single‐insert diploid parent is used. Gene‐centromere mapping of the cry3Aa transgene demonstrated that a non‐transgenic refuge might be naturally created in a TPS hybrid system through genetic recombination.     

GUS基因和NPTⅡ基因表达的相关性及其在转基因棉花检测研究中的应用

利用农杆菌介导转化法,将含有35S启动子驱动NPTII基因和GUS基因以及棉纤维特异表达启动子E6驱动目的基因FB的植物表达载体转入到常规棉花R15中.重点分析了GUS基因和NPTII基因在愈伤诱导阶段、T0代及T1代转基因棉花中的表达情况.综合两个基因的表达来进行转基因棉花的阳性鉴定,可以为转基因棉花后代的纯合选育提供双重保障.7个转基因株系选育到T3代共获得株行51个,卡那霉素检测多数株行阳性率在90%以上,其中21个株行阳性率达100%.     

Rht1(B. dw), an alternative allelic variant for breeding semi-dwarf wheat varieties

The vast majority of the world's acreage of semi-dwarf wheat varieties is at present cultivated with varieties carrying one of two genetically similar dwarfing genes, Rht1 and Rht2, derived from the Japanese variety ‘Norin 10’. Near-isogenic lines have been developed and tested to determine the breeding potential of an allelic variant of Rht1, designated Rht1(B. dw). Following its introduction into four varietal backgrounds, Rht1 (B. dw) was seen to reduce height by around 25%, to increase the number of grains setting in spikelets and ears by around 20%, to reduce grain weight by 10%, and to increase yields of plants grown under spaced or drilled conditions by about 8%. When compared to the commercially utilized Rht1 allele, as near isogenic lines in a ‘Mercia’ varietal background, Rht1 (B. dw) gives a significantly greater reduction in plant height, a greater increase in spikelet and ear fertilities, slightly less reduction in 1000-grain weight, and significantly higher spikelet, ear and plot yields. If these results are repeatable in other varietal backgrounds, over seasons, and under differing environmental conditions, Rht1 (B. dw) should have considerable commercial potential as an alternative allele for producing shorter-than-average, high-yielding, semi-dwarf wheat varieties.     

Pleiotropic effect of the dwarfing gene d18-k on cool tolerance at booting stage under the genetic background of an extremely cool-tolerant line Norin-PL8 in rice

Norin‐PL8 (‘PL8’) is an extremely cool‐tolerant line of rice in Japan that contains genes for cool tolerance originating from a javanica landrace. It was investigated to see whether the dwarfing gene d18‐k (kotaketamanishiki dwarf) exerts its pleiotropic effect on enhancing the cool tolerance at the booting stage in the genetic background of PL8. The d18‐k isogenic line of the recurrent parent PL8 (D8), PL8, and two commercial cultivars ‘Hayayuki’ and ‘Kirara 397’ were used. For each line/cultivar, the 12°C‐5‐day treatment was conducted at various times during the booting stage. In addition to spikelet fertility, the ratio (%) of the fertilized‐spikelet number of each treated panicle to the varietal mean of fertilized‐spikelet number per panicle in the control (FS‐T/C) was adopted to estimate cool temperature damage. For FS‐T/C, the lines‐cultivars ranked in the order of D8 > PL8 > ‘Hayayuki’ > ‘Kirara 397’, reflecting their cool tolerances. D8 exceeded PL8 in both pollen grain number per anther in the control and as an indicator of pollen fertility after the treatment, as a result of the effects of d18‐k. Consequently, d18‐k can be used to develop super‐highly cool‐tolerant cultivars for cool‐weather areas.     

Transgene introgression from Brassica napus to different varieties of Brassica juncea

Transgene introgression from transgenic rapeseed (Brassica napus) to different varieties of B. juncea was assessed in this study. Crossability between a transgenic rapeseed line Z7B10 (pollen donor) and 80 cultivars of 16 B. juncea varieties (including two wild accessions) was estimated by artificial pollination in a greenhouse. As a result, interspecific crossability between the transgenic Z7B10 line and the 80 B. juncea cultivars varied considerably, with seeds per flower from 0.00–10.67. Seed germination rates of the interspecific F₁ hybrids ranged from 49.0%–89.3%. The estimated frequencies of natural gene flow from the transgenic Z7B10 line to 10 B. juncea cultivars with different uses in the experiment field varied from 0.08% to 0.93%. The natural F₁ hybrids were highly sterile, with seeds per silique ranging from 0.27 to 1.03. In addition, seeds per flower of hybrid descendants varied from 0.02 to 0.22 when F₁ hybrids were self‐pollinated, and those ranged from 0.03 to 0.30 when F₁ hybrids were backcrossed with their corresponding B. juncea parents. Results of this study suggest a low level of transgene introgression from transgenic rapeseed to different B. juncea varieties, which provides a sound scientific basis for the safety management of coexisting transgenic B. napus and B. juncea varieties in China.     

RNAi‐based Bean golden mosaic virus‐resistant common bean (Embrapa 5.1) shows simple inheritance for both transgene and disease resistance

Bean golden mosaic virus (BGMV) is the causal agent of bean golden mosaic of common beans. A transgenic bean line that has been developed based on RNA interference to silence the BGMV rep gene showed immunity to the virus. Crosses were done between the transgenic line and six bean cultivars followed by four backcrosses to the commercial cultivars ‘Pérola’ and ‘BRS Pontal’. The transgene locus was consistently inherited from the crosses analysed in a Mendelian fashion in the segregating populations. The disease resistance reaction co‐inherited with the transgene. Nevertheless, the expression of disease resistance displayed a dosage effect phenomenon in the F₁ generation. The analysis of the homozygous near‐isogenic lines in field conditions, under high BGMV disease incidence, indicated that the transgenic lines were completely resistant. These results show the strength of the disease resistance obtained, the stability of the trait across generations and its usefulness in the management of a disease for which there is no reported Phaseolus germplasm with immunity.     

Greenhouse evaluation and inheritance analysis of transgenic cotton expressing <Emphasis Type="Italic">PTM3</Emphasis>, a MADS-box gene of Aspen

The PTM3 (MADS-box) gene characterized from flower of Aspen was ectopically expressed in cotton and was found to result in desirable agronomic traits. Among several transgenic lines, the PTM3 cotton event-10 was found to flower significantly earlier than the control and yield better [Ramachandran et al. (2011)]. We report our findings on performance based on greenhouse evaluation and inheritance of PTM3 cotton event-10. The T₁ progeny of event-10 were grown in pots under RBD design. The progeny were confirmed by kanamycin imbibition test, PCR, and GUS assay which showed the segregation of transgene at about 3:1 ratio. GUS assay was performed with pollen from all transgenic plants; plants that expressed gus in all pollen versus those that showed segregation for expression were found to be at a ratio of 1:2. Similar to observations made in the T₀ generation of event-10, the agronomic evaluation of T₁ progeny exhibited, on an average, earliness of 13 days in flowering, 13.5 days in crop maturity, and 22% of yield enhancement. The T₂ progeny of homozygous lines were grown on field soil in the greenhouse under strip trial design to see the effect as observed from potted plants of T₁ progeny. Inheritance of transgene cassette to T₂ progeny was confirmed by the same tests used to analyze T₁ progeny. As exhibited by T₀ and T₁ progenies, the T₂ progeny also showed earliness of 8 days in flowering, 12 days in crop maturity, and 17% of yield enhancement. The data generated from the progeny over two generations confirms that the PTM3 cotton event-10 is superior in agronomic characters as compared to non-transgenic cotton and is of interest for breeding shorter duration varieties with improved yield.     

Transgenic barley by particle bombardment. Inheritance of the transferred gene and characteristics of transgenic barley plants

Summary Transgenic barley plants (Hordeum vulgare L. cv. Kymppi) were obtained by particle bombardment of various tissues. Immature embryos and microspore-derived cultures were bombarded with gold particles coated with plasmid DNA carrying the gene coding for neomycin phosphotransferase II (NPTII), together with plasmid DNA containing the gene for -glucuronidase (GUS).Bombarded immature embryos were grown to plants without selection and NPTII activity was screened in small plantlets. One plant proved to be transgenic (T₀). This chimeric plant passed the transferred nptII gene to its T₁ progeny. The presence of the nptII gene was demonstrated by the PCR technique and enzyme activity was analyzed by an NPTII gel assay. Four T₀ spikes and 15 T₁ offspring were transgenic. The integration and inheritance was confirmed by Southern blot hybridization. Transgenic T₂ and T₃ plants were produced by isolating embryos from green grains of transgenic T₁ and T₂ plants, respectively and growing them to plants. After selfing, the ratio of transgenic to non-transgenic T₂ offspring was shown to follow the rule of Mendelian inheritance. The general performance of transgenic plants was normal and no reduction in fertility was observed.Microspore-derived cultures were bombarded one and four weeks after microspore isolation. After bombardment, cultures were grown either with or without antibiotic selection (geneticin ^R or kanamycin). When cultures were grown without selection and regenerated plants were transferred to kanamycin selection in rooting phase, one out of a total of about 1500 plants survived. This plant both carried and expressed the transferred nptII gene. The integration was confirmed by Southern blot hybridization. This plant was not fertile.     

Analysis on the spectra and level of wide-compatibility conferred by three neutral alleles in inter-subspecific hybrids of rice (Oryza sativa L.) using near isogenic lines

Wide‐compatibility varieties are a special class of rice germplasm that is able to produce fertile hybrids when crossed to either indica or japonica subspecies. Previous studies determined the f5 allele from ‘Dular’ (f5‐Du), f6 allele from ‘Dular’ (f6‐Du) and S5 allele from ‘02428’ (S5‐08) as neutral alleles conferring wide‐compatibility. However, the possible extent of the effect of the three neutral alleles has not been fully characterized because of the narrow range of the tester varieties used and the highly complex differentiation in Asian cultivated rice. In this study, we further developed the five near‐isogenic lines with higher recovery rates of the recurrent parent genome, and testcrossed to 14 japonica varieties, which have been widely used in rice breeding programmes in China. The results clearly revealed that all three neutral alleles exhibited substantial effects on spikelet fertility in most of the indica–japonica testcrosses, which indicated that these hybrid sterility loci have been extensively differentiated between indica and japonica varieties. The magnitudes of effects on spikelet fertility averaged over various crosses seem to be similar among the three neutral alleles, with f5‐Du, f6‐Du and S5‐08 alleles increasing spikelet fertility by 15.09%, 13.99% and 14.25%, respectively. The testcrosses involving f5‐Du allele generally showed much smaller variation in pollen fertility than others. The pyramiding lines with two neutral alleles showed a wider spectrum and a higher level of wide compatibility than others, whereas most of the increases in hybrid fertility couldn’t be simply explained by additive effects, suggesting the very complexity of wide compatibility and hybrid sterility. The indica–japonica hybrids involving restorer lines as one of their parents showed much higher pollen fertility (almost normal) and also higher spikelet fertility. The implications of the findings in rice breeding programmes are also discussed.     

Participatory farmers’ selection of common bean varieties (Phaseolus vulgaris L.) under different production constraints

On‐farm evaluation of 15 common bean varieties was undertaken with nine farmer groups under two fertilizer applications levels in four subcounties of Hoima and Rakai districts for two seasons to select farmers’ desired and undesired varieties. Farmers’ votes for acceptance and rejection of varieties at podding stage were converted to a preference index, and analysis of variance was conducted to examine differences in farmers’ preference indices among subcounties and combined across subcounties, seasons, management and gender. Management had no‐significant influence on the way farmers selected varieties in the subcounties. Choice of varieties varied significantly (p ≤ .001) between seasons and gender in the different subcounties. Variety Masindi Yellow Long and Farmers’ seed (Kaduli), and the introduced KATB1 were accepted by farmers due to their medium seed size, desired seed colour and potential of varietal adaptability to their farm conditions. Varieties NABE2, ROBA1 and RWR719 were deselected due to possession of traits less desired in the market. These results demonstrate the need for breeding programmes to involve diverse stakeholders in capturing the diverse traits preferences in varietal development process.     

A YA-type cytoplasmic male-sterile source in common wheat

A new cytoplasmic male‐sterile (CMS) system for hybrid wheat breeding, YA‐type CMS line with the cytoplasmic mutant from the common wheat variety ‘CA8057’, was developed by the Institute of Genetics and Developmental Biology, Chinese Academy of Sciences. The pollen sterility of YA‐type CMS line was easily maintained but difficult to restore. Some sterile lines with desirable agronomic performance, such as msYA‐‘CA8057’ (BC₁₇), msYA‐‘Yuandong 6’ (BC₉), msYA‐‘Jin 411’ (BC₉), msYA‐‘WL1’ (BC₁₀), msYA‐‘Yanshi 9’ (BC₁₀), msYA‐‘BPm16’ (BC₉), msYA‐‘Jindong 8’ (BC₉) and msYA‐‘Jinmai 33’ (BC₉), were bred and a restorer line GR1 was screened with 26 new restorer lines being developed by transferring restorer genes from GR1. It was found that abnormal phenomena occurred at the uninucleate‐pollen stage and the abortive pollen was poor in starch content and other components. The variance analysis of agronomic traits in eight sterile lines indicated that there was no general negative effect of cytoplasm. The genetic analysis for fertility restoration showed that two pairs of independent major genes (designated YARf₁YARf₁YArf₂YArf₂) and some minor genes could be involved in the fertility restoration in restorer line GR1, and YARf₁ was epistatic over YARf₂ for the genetic effect of fertility restoration. As a new CMS system, the YA‐type CMS line was of potential value for hybrid wheat breeding and should be further studied.     

Genetic analysis of resistance to green leafhopper, Nephotettix virescens (Distant), in three varieties of rice

The genetics of resistance to green leafhopper, Nephotettix virescens (Distant), in rice varieties ‘IR36’ and ‘Maddai Karuppan’ and breeding line ‘IR20965‐11‐3‐3’ was studied. The reactions of F₁ hybrids, F₂ populations and F₃ lines from the crosses of test varieties with the susceptible variety ‘TN1’ revealed that resistance in ‘IR36’ and ‘Maddai Karuppan’, is governed by single recessive genes while resistance in ‘IR20965‐11‐3‐3’ is controlled by a single dominant gene. Allele tests with the known genes for resistance to green leafhopper revealed that the recessive gene of ‘IR36’ is different from and inherited independently of Glh1, Glh2, Glh3, Glh4, Glh5, Glh8 and Glh9t. This gene is designated as glh10t. The recessive gene of ‘Maddai Karuppan’ and the dominant gene of ‘IR20965‐11‐3‐3’ are also non‐allelic to Glh1, Glh2, Glh3, Glh4, Glh5 and Glh8t. Thus, the dominant gene of IR20965‐11‐3‐3 is designated as Glh11t. The allelic relationships of the recessive gene of ‘Maddai Karuppan’ with glh8 and glh10t should be investigated.     

Relationship between rice grain amylose and pasting properties for breeding better quality rice varieties

One hundred and sixty‐seven rice varieties/laudraces from 23 different rice‐growing countries were used for studying the relationship between the amylose content (AC %) and paste viscosity parameters (RVU) through Pearson correlation. AC and paste viscosity parameters were measured using Auto Analyser and Rapid Visco Analysers respectively. Based on all five RVA pasting properties, i.e. peak viscosity (PV), minimum viscosity (MV), breakdown (BD), final viscosity (FV) and setback (SB), the simple paired t‐test (at P = 0.05) was carried out to identify varieties that showed non‐significant differences (similar) from the check variety, separately for each of the four AC groups, e.g. Waxy rice AC (0–8%, n = 21) –‘IR29’ (Philippines); Low AC (8.1–16%, n = 58) –‘Koshihikari’ (Japan); Intermediate AC (16.1–24%, n = 64) –‘Local Sadri’ (Iran); high AC (>24.1%, n = 24) –‘Kasalath’ (India). Significant variation among the rice varieties for AC and pasting properties were observed. Means of the AC and the viscosity parameters including PV, MV, BD, FV and SB were 15.8, 416.1, 180.3, 235.8, 380.9 and 200.6 RVU respectively. Significant correlations existed between AC and paste viscosity parameters at AC group level. The paste viscosity parameters, i.e. PV, BD and SB are important within an AC group as far as cooking quality is concerned. In the low AC group, ‘Koshihikari’ (check) was most preferred by Japanese consumers, which was used for identifying 16 varieties that showed non‐significant differences for pasting properties with it. Some of the important Japanese varieties were ‘Todorokiwase’, ‘Owarihatamochi’, ‘Sasanishiki’, ‘Reimei’ and ‘Kinuhikari’. These varieties had good cooking quality with low AC, high PV and BD with low SB. In contrast, the Iranian cultivar ‘Local Sadri’ with good cooking quality had intermediate levels of AC and SB while recording high PV and BD. The varietal diversity for pasting properties available within each AC group may allow the breeder to make effective selections for improving the cooking quality according to consumer choice.     

Transgenic soybean with low phytate content constructed by <Emphasis Type="Italic">Agrobacterium</Emphasis> transformation and pollen-tube pathway

The expression of a microbial phytase in transgenic plants may create a new biochemical pathway that mobilizes its endogenous phytate and release inorganic phosphate from it, so that more phosphorus is available for plant growth. In this study, transgenic soybean plants were generated via both Agrobacterium transformation and pollen tube pathway with the PhyA gene of Aspergillus ficuum. The optimal concentrations of plant hormones including N₆-benzylaminopurine (BAP), gibberellin (GA₃) and indole-3-butyric acid (IBA) were tested based on their effectiveness on promoting the growth of transgenic explants. Genomic PCR results and Southern blot hybridization analysis showed that transgenic soybean plants selected for resistance to kanamycin contained the phyA transgene. The transgenic soybean plants with phyA gene integrated in their genome exhibited lower amount of phytate in different soybean tissues including leaf, stem and root, which indicated that engineering crop plants with a higher expression level of heterologous phytase could improve the degradation of phytate and potentially in turn mobilize more inorganic phosphate from phytate and thus reduce phosphate load on agricultural ecosystems.     

Role of male-sterile cytoplasm in resistance to barley yellow mosaic virus and Fusarium head blight in barley

Two types of male‐sterile cytoplasm, designated msm1 and msm2, in barley were investigated to determine whether these cytoplasms confer resistance to barley yellow mosaic virus (Ba YMV) and Fusarium head blight (FHB). Alloplasmic lines and isogenic lines of two cultivars showed the same reaction to each Ba YMV as that of their euplasmic lines. This demonstrates that the barley male‐sterile cytoplasms msm1 and msm2 have no effect on resistance to BaYMV. No significant difference in reactions to FHB was recognized among fertile alloplasmic lines of ‘Adorra’, but the difference in reactions to FHB between fertile and sterile isogenic lines of ‘Adorra’ was significant. The damage caused by FHB in the male‐sterile lines that produced sterile pollen was significantly greater than the damage in a sterile line that did not produce pollen. These results suggest that pollen or anthers are important factors in infection with or spread of FHB. For production of hybrid seeds, male‐sterile lines with no pollen production, such as those with msm1 male‐sterile cytoplasm, would reduce FHB infestation.     

Evaluation of cultivated barley (Hordeum vulgare) germplasm for the presence of thermostable alleles of β-amylase

A cleaved amplified polymorphic sequence marker was used to detect the alleles Bmy‐Sd2H and Bmy‐Sd3 identifying highly thermostable isoforms of the enzyme b‐amylase, which improves fermentability during brewing. Among the 889 accessions of barley (Hordeum vulgare) investigated, and two accessions of H. spontaneum a total of 166 accessions were identified carrying the superior b‐amylase alleles. These thermostable alleles of b‐amylase were most frequently observed in six‐rowed varieties originating from Asia, especially Japan, with 61.9% of the accessions from Asia carrying the alleles of interest. Additional six‐rowed barleys carrying the relevant alleles were identified among accessions from America, Africa and the Near East. In the European varieties, the percentage of accessions with the alleles of interest was 5.1% with a strong predominance in two‐rowed spring barleys. A pedigree analysis identified the cross ‘Binder’ x ‘Gull’ as the main source of the thermostable b‐amylase alleles in European varieties. The data suggest that an improvement of malting quality in barley could be achieved by introduction of the Bmy1‐Sd2H and Bmy1‐Sd3 alleles into the European breeding programmes.     

Marker‐assisted backcross breeding for improvement of drought tolerance in bread wheat (Triticum aestivum L. em Thell)

Drought stress is presently a major productivity limiting factor in wheat. This study developed five wheat lines with inbuilt tolerance to drought stress using marker‐assisted backcross breeding (MABB) approach employing three linked quantitative trait loci (QTLs) in an initial population of 516 BC₁F₁ plants. The high‐yielding wheat cultivar ‘HD2733’ grown over last few years extensively in the eastern plains of India is largely sensitive to drought and is used as the recurrent parent. ‘HI1500’ released for water‐limiting conditions and carrying drought‐tolerant QTLs was used as donor parent. MABB lines were advanced using foreground and background selection, coupled with stringent phenotyping. We identified 29 lines that were homozygous for targeted QTLs in different combinations with background recovery range of 89.2%–95.4%. Further evaluation of selected lines for physiological traits and distinctness, uniformity and stability (DUS) characters under rainfed condition identified five potential varieties for national varietal evaluation programme in the zone. The report is first of its kind in implementing known QTLs for the development of drought‐tolerant wheat lines through MABB approach.     

Genetic Structure of Three Open-Pollinated Faba Bean Varieties (Vicia faba L.)*

To study the genetic structure of open-pollinated faba bean varieties, three sets of genotypes were generated from each of the varieties ‘Minica’, ‘Kristall’, and ‘Deiniol’: (1) inbred lines developed by single-seed descent in bee-proof isolation cages, (2) intravanetal and (3) intervarietal F₁-hybrids produced by controlled hand crossing. In 1989 and 1990, a total of 144 entries, including the three open-pollinated source varieties, were grown in single-row plots with two replications at two locations in Western Germany. Performance data of the inbred lines revealed relatively large genotypic variability within each variety, for all the characters studied, which was greater in ‘Deiniol’ and ‘Kristall’ than in ‘Minica’. An average yield heterosis of 20 %, 39 % and 57 %, respectively, was found for the intravarietal hybrids, whereas that of the intervarietal hybrids varied between 70 and 73 %. Compared to the mean yield of the inbred lines the superiority of the open-pollinated source variety was small and not significant in ‘Minica’ (6 %), but it was large and highly significant in ‘Kristall’ (22 %) and ‘Deiniol’ (37 %). The present investigation revealed in faba bean that similar performance levels of open-pollinated varieties may result from either high per se performance under inbreeding with low heterosis values or from lower per se performance but with high heterosis values.     

Limitations of PCR‐based molecular markers to identify male‐sterile and maintainer plants from Indian onion (Allium cepa L.) populations

The main aim of this study was to validate PCR markers for determining cytoplasm and genotypes at the Ms locus in short‐day onion. Three cytoplasmic (OSN, MKFR and accD) and four nuclear (OPT, jnurf13, AcSKP1 and AcPMS1) markers were employed. Sel. 121‐1 had 100% S cytoplasm, whereas Sel. 121‐2, ‘Pusa Red’ and ‘Pusa Madhavi’ had 88%, 33% and 17% S cytoplasm, respectively. ‘Early Grano’ and ‘Pusa Riddhi’ did not possess S cytoplasm. Observations in 33 commercial varieties revealed two with sterile (S) cytoplasm. Nuclear markers were not found in linkage disequilibrium with the Ms locus, and the constitution of Ms alleles by OPT was different from other three markers, which were in conformity with each other. The other three markers predicted that most of the plants should be homozygous recessive. Anther colour also did not confirm the sterility status. It can be concluded that accD may be used for cytoplasm determination based on the ease of its use. For the Ms locus tagging, more markers are needed to be evaluated to isolate maintainer lines from open‐pollinated populations.     

A cytogenetic study of bilateral sexual polyploidization in cassava (Manihot esculenta Crantz)

Microsporogenesis and megasporogenesis were cytogenetically and histologically analysed in three cassava clones:‘Rayong 1′,‘Rayong 60′,‘M. mga’ and two hybrid lines,‘OMR 3641‐1’ and ‘OMR 3641‐1’ to elucidate the evolution of sexual polyploids in cassava. At telophase II, formation of 17‐21 micronuclei per pollen cell plate was observed in 16 out of 351 cell plates in ‘M. mga’. Micronuclei were observed at low (0.3‐2.3%) frequencies, at the sporad stage in all clones. Monads, dyads, triads and tetrads, which are established sources of high ploidy levels were observed at low (2.6%) and high (22.2%) frequencies. Megasporogenesis in ‘Rayong 1’ and ‘Rayong 60’ showed a lack of second meiotic divisions after a successful first division that resulted in partly unreduced embryo sacs with 2n eggs, suggesting another unrecognized and, as yet, unreported source of sexual polyploid formation in cassava. Meiotic abnormalities during microsporogenesis and megasporogenesis are implicated as being responsible for the formation of mixoploids (triploids and tetraploids) in cassava breeding programmes. A cytogenetic mechanism resulting in bilateral sexual polyploids through different gametic fertilization pathways in cassava is suggested and its role in breeding is briefly discussed.