The effect of starch and ineubation temperature in anther culture of potato

Three Andean tetraploid potato genotypes (2n=48) and 7 anther-derived dihaploids (2n=24) originating from two of the tetraploids were used in anther culture. Relative number of embryos/vial was significantly higher when the anther culture media was gelatinized with 3% potato starch than when Gelrite or wheat starch (3%) were used as gelatinizing agents. The degree of anther culture response varied between tetraploids but also within a group of related dihaploids. Additionally, the embryo production of individual genotypes, tetraploids as well as dihaploids, was dependent on the incubation temperature (10, 15, 20, 25, 30°C) of the anther culture. The incubation temperature of the anther culture was also important for the regeneration rate. Direct regeneration was mostly stimulated when the anther culture was incubated at 20°C.Abbreviations BAP 6-benzylaminopurine - IAA indole-3-acetic acid     

Genetical analysis of in vitro cell and tissue culture response in potato

The genetics of tissue culture response in potato has been examined by analysing a sample of dihaploids (2n=2x=24) extracted from tetraploid parents (4n=4x=48). The genotypes were screened for rate of nodal multiplication, in vitro tuberisation, regeneration from leaf discs and protoplast plating efficiency. Significant differences were detected between dihaploids for the traits measured and this indicates that tissue culture response in the tetraploid parents must be in the heterozygous condition. Estimates of the broad sense heritabilities were calculated together with the number of genes or effective factors involved in the control of the traits. These estimates indicate that tissue culture response in potato is under relatively simple genetic control and blocks of genes may be located on specific chromosomes. The inheritance of RFLP markers in the segregating dihaploid population was also monitored and the potential of using molecular markers linked to gene(s) controlling tissue culture response is discussed.     

Phenotypic characterization and bulk segregant analysis of anther culture response in two backcross families of diploid potato

Two diploid (2n=2x=24) backcross potato populations (PBCp, and CBC) were characterized for anther culture response (ACR). PBCp (Solanum phureja Juz. & Buk. genotype 1-3 × CP2) and CBC (CP2 × S. chacoense Bitt. genotype 80-1) resulted from a cross between CP2 (intermediate ACR) and its parents, S. chacoense 80-1(low ACR) and S. phureja 1-3 (high ACR). Three components of ACR were initially investigated: embryos per anther (EPA), embryo regeneration rate and percent monoploids (2n=1x=12) among regenerants. EPA was selected for further characterization because of its relative stability. In a series of studies of EPA on a total of 44 genotypes within CBC, nine high (mean EPA=2.5) and ten low (mean EPA=0.02) selections were made. In PBCp, ten high (mean EPA= 4.7) and ten low (mean EPA= 0.05) selections were made from 67 genotypes. High and low selections were used for bulk segregant analysis to screen 214 RAPD primers as candidate markers linked to EPA. Bands amplified by OPQ-10 and OPZ-4 were associated in coupling and repulsion, respectively, to ACR in PBCp. A band amplified by OPW-14 primer was associated in coupling to ACR in CBC. One-way ANOVAs using presence/absence of each candidate band to classify additional genotypes in each population verified association of the markers with EPA.     

QTL analysis of potato tuberization

Quantitative trait loci (QTLs) affecting tuberization were detected in reciprocal backcrosses between Solanum tuberosum and S. berthaultii. Linkage analyses were performed between traits and RFLP alleles segregating from both the hybrid and the recurrent parent using a set of framework markers from the potato map. Eleven distinct loci on seven chromosomes were associated with variation in tuberization. Most of the loci had small effects, but a QTL explaining 27% of the variance was found on chromosome 5. More QTLs were detected while following alleles segregating from the recurrent S. tuberosum parent used to make the backcross than were detected by following alleles segregating from the hybrid parent. More than half of the alleles favoring tuberization were at least partly dominant. Tuberization was favored by an allele from S. berthaultii at 3 of the 5 QTLs detected by segregation from the hybrid parent. The additive effects of the QTLs for tuberization explained up to 53% of the phenotypic variance, and inclusion of epistatic effects increased this figure to 60%. The most common form of epistasis was that in which presence of an allele at each of 2 loci favoring tuberization was no more effective than the presence of a favorable allele at 1 of the 2 loci. The QTLs detected for tuberization traits are discussed in relationship to those previously detected for trichome-mediated insect resistance derived from the unadapted wild species.Paper number 54 of the Department of Fruit and Vegetable Science, Cornell University     

QTL analysis of potato tuber dormancy

The potential loss of chemical sprout inhibitors because of public concern over the use of pesticides underscores the desirability of breeding for long dormancy of potato (Solanum tuberosum L.) tubers. Quantitative trait locus (QTL) analyses were performed in reciprocal backcrosses between S. tuberosum and S. berthaultii toward defining the complexity of dormancy. S. berthaultii is a wild Bolivian species characterized by a short-day requirement for tuberization, long tuber dormancy, and resistance to several insect pests. RFLP alleles segregating from the recurrent parents as well as from the interspecific hybrid were monitored in two segregating progenies. We detected QTLs on nine chromosomes that affected tuber dormancy, either alone or through epistatic interactions. Alleles from the wild parent promoted dormancy, with the largest effect at a QTL on chromosome 2. Long dormancy appeared to be recessive in the backcross to S. berthaultii (BCB). In BCB the additive effects of dormancy QTLs accounted for 48% of the measured phenotypic variance, and adding epistatic effects to the model explained only 4% more. In contrast, additive effects explained only 16% of the variance in the backcross to S. tuberosum (BCT), and an additional 24% was explained by the inclusion of epistatic effects. In BCB variation at all QTLs detected was associated with RFLP alleles segregating from the hybrid parent; in BCT all QTLs except for two found through epistasis were detected through RFLP alleles segregating from the recurrent parent. At least three dormancy QTLs mapped to markers previously found to be associated with tuberization in these crosses.Paper number 55 of the Department of Fruit and Vegetable Science, Cornell University     

Influence of 2,4-d and lactose on pollen embryogenesis in anther culture of potato

Anthers of diploid genotypes of Solanum tuberosum capable of androgenesis were cultured on different media to examine the effect on induction of pollen embryogenesis of 2,4-d and lactose. Anthers cultured in callogenic medium with 2,4-d and sucrose produced pollen derived embryoids only exceptionally. When sucrose was replaced by lactose the frequency of embryogenesis was as high or higher than in embryogenic auxin-free medium. Substitution of lactose for sucrose in the embryogenic medium had no effect. Supplementing the embryogenic medium with 2,4-d strongly reduced the frequency of pollen embryoids in the presence of sucrose but not with lactose.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid     

Barley anther culture using membrane rafts

When compared to agarose solidified media in small petri dishes, membrane rafts used in conjunction with liquid induction media significantly improved anther culture response in the Australian, malting-quality, spring barley cultivar Clipper. In contrast, the German cultivar Gimpel did not show an increased response on rafts.Abbreviations BA 6-benzylaminopurine - IAA indoleacetic acid - DH doubled haploid     

The influence of culture conditions on anther culture response of commercial varieties of Solanum tuberosum L.

The effect of media manipulatioss, temperature pretreatment, carbohydrate source, and seasonal variation on tetraploid potato anther cultures was investigated. The anther culture responses of three commercial Nordic potato varieties from Scandinavia and two from Germany were compared on different media manipulations. With most of the varieties, solid MS media gave better yields than other published media manipulations. Pretreatments at +6°C and at +30°C were studied on Pito and Danva varieties. The +6°C pretreatment and no pretreatment had the same effect on the anther culture response of cv. Pito, while with cv. Danva pretreatment at +6°C promoted embryogenesis. The +30°C pretreatment had no positive effect on anther culture response on either cultivar. The effect of maltose, melibiose and mannitol individually and in combination with sucrose were compared to normal sucrose medium in cv. Pito anther cultures. Anthers incubated on normal sucrose medium gave the highest embryoid and plant yields; the second highest plant yields were obtained on pure maltose medium. Strong seasonal variation was observed throughout the year in cv. Pito anther cultures. The percentage of anthers producing embryoids ranged from 15–20% during September and October to just 1–3% from February through May. The annual average embryoid production rate was 6.18%.     

Effect of maltose on the response of potato anthers in culture

Anthers of the Solanum tuberosum genotype H3703 were cultured on medium containing equimolar concentrations of sucrose or maltose. It was found that significantly more pollen embryos became plants after culture on maltose and hence the yield of plants per 100 anthers cultured increased significantly. Mechanisms by which carbohydrate source may influence response to anther culture are discussed.     

The potato in Spain during the late 16th century

A study is presented of the Hospital de la Sangre account books in Seville at the Archivo Hispalense for the period 1546 to 1601, to verify purchases of potatoes (Solanum tuberosum) during that period. Potatoes were bought regularly in the Seville market from 1580 onwards, with the first record appearing in 1573 thus agreeing with Salaman’s conclusion that potatoes became established in Spain by about 1570. Purchases were confined almost entirely to December and January each year, lending weight to the hypothesis that these were short day adaptedS. tuberosum ssp.andigena, actually grown in Spain and forming tubers in the short days at the end of the year. If potatoes had been imported for direct consumption from South America shipping records indicate that they could have arrived in Seville at all times of the year. A listing of other fruits and vegetables bought in the Seville market shows a wide range of mostly mediterranean crops with some of Near Eastern origin and certain spices imported through Lisbon from the Ear East. Surprisingly, very few New World crops are mentioned.     

An efficient genotype-independent method for regeneration of potato plants from leaf tissue

The regeneration of plants from leaf explants of a number of potato cultivars using a number of published one-, two- and three-step methods was assessed. A method using a pretreatment with high levels of auxin and cytokinin coupled with silver thiosulphate in the regeneration medium proved the most rapid and efficient for the eight cultivars examined.Abbreviations BA benzyladenine - GA₃ gibberellic acid - IAA indoleacetic acid - MS Murashige and Skoog - NAA naphthaleneacetic acid - STS silver thiosulphate     

Restriction fragment length polymorphism (RFLP) analyses of plants produced by in vitro anther culture of Solanum chacoense Bitt

Summary Green mesophyll protoplasts of the dihaploid potato line 1982 (Solanum tuberosum L.) were fused with herbicide-bleached mesophyll protoplasts of the dihaploid potato line 679 using a polyethylene glycol protocol. Heterokaryons were identified under a fluorescence microscope using the dual fluorescence of carboxyfluorescein-stained, herbicide-bleached protoplasts and the autofluorescence of green mesophyll protoplasts. About 20% of the protoplasts survived the fusion treatment, and the fusion frequency was 3%–4%. Unfused and fused protoplasts were mass cultured for 6 weeks after which vigorously growing calli were selected and transferred to shoot regeneration medium. Somatic hybrids were identified by a combination of five isozyme markers, and the ploidy level was determined by flow cytometry. Out of 15 calli that regenerated shoots, 6 plants derived from 2 different calli were identified as hexaploid somatic hybrids, while one morphologically deviant plant from a third callus was identified as a mixoploid that had lost some enzyme markers after 4 months of culturing.     

Characterisation of the S-adenosylmethionine decarboxylase (SAMDC) gene of potato

S-adenosylmethionine decarboxylase (SAMDC) is involved in the biosynthesis of the polyamines, spermidine and spermine. Recently, we reported the isolation of a putative cDNA clone of the SAMDC clone of potato (Plant Mol Biol 20; 641–651). In order to confirm that the potato genes does encode SAMDC, a complementation experiment with a yeast strain that possesses a null mutation in the SAMDC gene was performed. The yeast strain contains a deletion-insertion mutation in the SAMDC gene and has an absolute requirement for the addition of exogenous spermidine for growth. When the full-length potato cDNA was expressed in the mutant yeast strain there was no longer a requirement for exogenous spermidine. Immunoblotting experiments suggest that the potato SAMDC gene product has an apparent molecular mass of 39 kDa. Expression of the SAMDC gene was high in the young and actively dividing tissues and low in the mature and non-dividing tissues of both vegetative and reproductive organs. Additionally, isolation and characterisation of the corresponding genomic clone is reported. The gene has one intron in its 5-untranslated sequence but otherwise the transcribed portion is identical to the cDNA clone.     

A comparison of several published methods for barley anther culture

A number of methods have been published for barley (Hordeum vulgare L.) anther culture and have gained acceptance in different laboratories. The breeder's requirement is for a compromise method that gives good, repeatable results for a wide range of genotypes. Yet the routine production of spontaneously doubled haploid green regenerants remains difficult. Despite attempts to formulate a widely-applicable anther culture method, the 4 main published methods, compared here with one modified procedure, are quite distinct for a number of important characteristics. The methods interacted strongly with the 3 genotypes, and response ranged from zero to 28 green regenerants per 100 anthers plated. The current methods still require often substantial modification to suit local situations in order that the technology may be exploited by barley breeders.Abbreviations BAP benzylaminopurine - DH doubled haploid - FV final volume - IAA indoleacetic acid - IBA indole-3-butyric acid - MS Murashige & Skoog - PABA para-aminobenzoic acid     

Initiation and culture of potato tuber callus tissue with picloram

Callus cultures of 7 potato cultivars were initiated from tuber tissue and maintained on Gelrite-solidified media with 1–20 M picloram as the only PGR. Ten M picloram was the optimal concentration for callus induction. By 4–6 weeks after explanting, there was sufficient callus produced for subculture to maintenance media which contained 1–20 M picloram as the only PGR. When grown in the dark at 25°C, subcultured callus typically increased 10-fold in wet weight in 4–5 weeks. The callus produced was friable and a light grey to cream color. Callus cultures were used to establish cell suspension cultures. Callus and cell suspension cultures have been maintained for over 2 years on the picloram containing media.Abbreviations BA benzyladenine - 2,4-D 2,4-dichlorophenoxyacetic acid - MS Murashige-Skoog - NAA naphthaleneacetic acid - PGR plant growth regulator Research paper #9053 of the Idaho Agricultural Experiment Station.     

Anther culture of Solanum genotypes

Anther culture response was examined in five Solanum genotypes. Large genotypic differences exist in response to culture and liquid culture media were found to be superior to agar solidified media systems. Pre-treatment effects on embryoid induction were also investigated and two of the genotypes displayed differential response to temperature stress pretreatment. In general the beneficial effect of charcoal in the media was confirmed. Successful embryoid production and plantlet regeneration was obtained from the wild potato species, Solanum papita. The influence of sucrose concentration on embryoid production was also investigated. Large genotype by sucrose interactions were detected and this was mainly due to the differential response of the tuberosum clones to increasing sucrose concentration when compared with S. papita. Embryoids were produced from this species in media containing 15% sucrose although the optimum concentration of sucrose for embryoid production was 9%. The possible role of anther culture techniques in gene introgression and potato improvement is discussed.     

Gibberellic acid regulation of adventitious shoot formation from tuber discs of potato

Summary The formation of adventitious shoots from potato tuber discs explanted onto a modified Murashige and Skoog (MS) medium containingN ⁶-benzylaminopurine (BAP) (3.0 mg/l), and α-naphthaleneacetic, acid (NAA) (0.01 mg/l), was affected by gibberellic acid (GA). The presence of GA in the explant medium was required for shoot formation and 3×10⁻¹⁰ M GA appeared optimum. However, microscopic examination of the tissue protuberances on the surface of the tuber discs from which shoots arose revealed that GA inhibited the formation of shoot meristems. Tuber discs cultured for 6 wk on MS medium containing BAP and NAA without GA did not initiate adventitious shoots that could be determined visually, but microscopic examination of the tissue protuberances revealed the presence of numerous shoot meristems. Subsequent transfer of these tuber discs to medium with GA but without BAP or NAA resulted in the formation of shoots from 100% of the recultrued dises. Thus it appears that although GA inhibits shoot meristem initiation from potato tuber discs, it is required for shoot development once meristems are initiated. This is Journal Paper 8297 of the Purdue University Agricultural Experiment Station. The research was supported by Purdue University Agricultural Experiment Station Program Improvement Funds. Potato tubers were supplied by Wm. Gehring Farms, Inc., Rensselaer, Indiana.     

The use of cross prediction methods in a practical potato breeding programme

Summary Most previous studies on cross prediction methods have examined relatively few crosses, particularly in relation to the numbers involved in most breeding programmes. In this paper the feasibility of using cross prediction methods was examined in a practical potato (Solanum tuberosum) breeding scheme by the analyses of progeny from 52 crosses. The variate considered was breeder's preference, a visual assessment made of the harvested tubers to estimate their commercial potential. The results showed that it was possible to identify the superior crosses. Cross prediction based simply on the mean preference scores, averaged over scorers and clones within progenies, estimated on seedlings or first clonal year plants, provided the best estimate of a progeny's performance in the third clonal generation. Predictions based on the expected proportion of clones that would transgress a given target value also provided a good indication of a progeny's potential. The poorest prediction was obtained by using the observed frequency of desirable clones in a progeny sample. The implications for potato breeding are discussed.