Shoot and root fresh and dry weight, shoot length, leaf area, leaf area index and relative growth rate of 14 day old tepary bean (Phaseolus acutifolis Gray) and navy bean (Phaseolus vulgaris L.) seedlings were reduced following treatment with NaCl solution exhibiting osmotic potential of either -0.25, 0.50, and -0.75 MPa. Salinity reduced the growth of navy bean more than tepary bean. The physiological basis of the adaptive response of tepary bean seedlings to salt stress was explored by determining the water and osmotic potentials, relative water content, free amino acid and sugar concentrations, distribution and levels of inorganic ions within the seedlings and ATPase activity of the root plasma membrane. Salinity led to an osmotic adjustment in the leaves and the proximal part of the root of tepary bean. Turgor remained almost constant whereas osmotic and water potential and relative water content declined following the salt treatments. The osmotic adjustment of the leaves and proximal part of the roots was -1.7 MPa and -1.2 MPa, respectively, in seedlings treated with -0.75 MPa NaCl solution. Free amino acids and sugars increased under salinity stress in both species but they increased more in the tepary bean. Glucose was the most abundant free sugar. The nonstructural carbon solutes contributed -0.15 MPa to the seedling's osmotic adjustment whereas Na, Cl, K and Ca ion levels contributed -0.85 MPa. However, the levels of these solutes were not large enough to account for the total osmotic adjustment observed in the salt treated seedlings. This study shows that tepary bean has specific strategies to overcome the impact of salinity through osmotic adjustment and exclusion of Na and Cl ions from the stems and leaves by retaining these ions in the proximal part of root and stem base. (Abstract shortened with permission of author.)

Tissue culture and radicle excision techniques were employed to evaluate salt tolerance in alfalfa (Medicago sativa L.). Plant suspension cultures of either seedling root or shoot origin were studied in media with or without supplemental NaCl (3.54 g liter⁻¹). In most cases, the growth rates of root-derived cultures were stimulated by this low level of supplemental NaCl while most shoot-derived cultures were not stimulated by NaCl. Excised radicles of three populations of alfalfa which possessed widely differing ranges of germination salt tolerance were screened in four salts (NaCl, KCl, Na₂SO₄, and K₂SO₄) at six varying concentrations. As was observed in the tissue culture experiments, low levels of NaCl (7.09 g liter⁻¹) stimulated radicle elongation of all populations as compared to the elongation levels of the control solutions (no supplemental salts). In general, for NaCl, the population that posessed the highest degree of germination salt tolerance (Az-St 1982) also displayed the greatest rates of radicle elongation especially in the highest salt concentrations. Additionally, this population along with the moderately germination salt tolerant population (Az-ST 1979) maintained higher rates of elongation in KCl, K₂SO₄ and Na₂SO₄ than did the control germplasm which has little germination salt tolerance (Mesa Sirsa Control). Examinations of each individual population in all four salts simultaneously, indicated that the sulfate salts reduced radicle elongation to a greater extent than did the chloride salts. Evaluation of both osmotic effects and specific ion effects showed that the specific ion effects attributed to the anions were more detrimental to radicle elongation than were the osmotic effects.

Thesis (Ph.D.)--University of Hawaii at Manoa, 1994. / Includes bibliographical references (leaves 97-114). / Microfiche. / xiii, 114 leaves, bound ill. 29 cm

Thesis (Ph.D.)--Flinders University, School of Biological Sciences. / Typescript bound. Includes bibliographical references. Also available online.

Thesis (MSc.)--University of Stellenbosch, 2000. / ENGLISH ABSTRACT: The detrimental effect of soil salinity on crop production is a growmg problem worldwide (Tanji, 1990b). The degree to which plants can tolerate high concentrations of salt in their rooting medium is under genetic control with different genetic and physiological mechanisms contributing to salt tolerance at different developmental stages (Epstein & Rains, 1987). Only limited variation exists for salt tolerance in the cultivated cereals. This has prompted attempts to select tolerant progeny following hybridisation of cultivated species and wild, salt-tolerant species. Thinopyrum distichum, an indigenous wheatgrass that is naturally adapted to saline environments (McGuire & Dvorak, 1981), was crossed with triticale (x Triticosecale) in an attempt to transfer its salt tolerance and other hardiness characteristics (Marais & Marais, 1998). The aims of this study were to (i) identify Thinopyrum chromosomes carrying genes for salt tolerance and to identify molecular markers for these chromosomes, (ii) identify a number of diverse monosomic and disomie addition plants. Bulked segregant analysis (BSA), in combination with AFLP, RAPD and DAF marker analysis was implemented to screen for polymorphisms associated with salt tolerance. Five putative AFLP markers and two RAPD markers were detected using bulks composed of salt tolerant plants and bulks composed of salt sensitive plants. The distribution of the markers in these bulks suggests that more than one Thinopyrum chromosome carry genes for salt tolerance. Salt tolerant monosomic and disomie addition plants were characterised for AFLP, RAPD and DAF polymorphisms in an attempt to find markers associated with the chromosome(s) conditioning salt tolerance. One salt tolerant monosomic and one disomie addition plant was identified. One AFLP and two RAPD markers were identified for the Thinopyrum chromosome( s) present in the monosomic addition plant, while three AFLP and three RAPD markers were identified for the disomie addition plant. An attempt was also made to identify diverse chromosome addition plants having complete or near complete triticale genomes plus an additional random Thinopyrum chromosome. Plants with 2n = 43 /44 were identified and characterised for molecular markers (AFLP and RAPD). Cluster analysis was used to group the putative monosomic or disomie addition plants according to the specific Thinopyrum chromosomes they retained. Seventeen AFLP and RAPD markers could be used to group the 24 putative addition plants into six broadly similar groups with different additional Thinopyrum chromosomes. While the members of each group are likely to carry the same additional Thinopyrum chromosomes, this may not necessarily be the case as the interpretation of the marker results is complicated by heterogeneity among plants with regard to the triticale background chromosomes they possess. It is also likely that chromosome translocations occurred during backerossing which may further complicate data. Nonetheless, it is now possible to select disomie addition plants from each group that are likely to represent different Thinopyrum chromosomes. The data will also be useful in future attempts to find further addition plants carrying the remaining Thinopyrum chromosomes. / AFRIKAANSE OPSOMMING: Die skadelike effek van grond versouting op gewasproduksie neem wêreldwyd toe (Tanji, 1990b). Die mate waartoe plante hoë konsentrasies sout in die wortelstelsel kan hanteer is onder genetiese beheer en verskillende genetiese en fisiologiese meganismes dra by tot die soutverdraagsaamheid tydens verskillende ontwikkelingstadia (Epstein & Rains, 1987). Slegs beperkte variasie bestaan vir soutverdraagsaamheid in verboude grane. Dit het aanleiding gegee tot pogings om soutverdraagsame nageslag te selekteer na hibridisasie van verboude spesies en wilde, soutverdraagsame spesies. Thinopyrum distichum, 'n inheemse koringgras, wat aangepas is by brak omgewings (McGuire & Dvorak, 1981), is met korog (x Triticosecale) gekruis in 'n poging om die gene vir soutverdraagsaamheid en ander gehardheidseienskappe oor te dra (Marais & Marais, 1998). Die oogmerke van hierdie studie was om (i) Thinopyrum chromosome te identifiseer wat gene bevat vir soutverdraagsaamheid en molekulêre merkers te vind vir hierdie chromosome, (ii) 'n aantal diverse monosomiese en disomiese addisieplante te identifiseer. Bulksegregaatanalise (BSA), gekombineer met AFLP-, RAPD- en DAF-merkeranalise, is gebruik om polimorfismes geassosieerd met soutverdraagsaamheid op te spoor. Vyf moontlike AFLPmerkers en twee RAPD-merkers is geïdentifiseer met gebruik van bulks bestaande uit soutverdraagsame plante en bulks bestaande uit soutgevoelige plante. Die verspreiding van die merkers in soutverdraagsame bulks dui daarop dat meer as een Thinopyrum chromosoom bydra tot soutverdraagsaamheid. Soutverdraagsame, monosomiese en disomiese addisieplante is gekarakteriseer vir AFLP- en RAPD-polimorfismes in 'n verdere poging om merkers te vind vir chromosome betrokke by soutverdraagsaamheid. Een soutverdraagsame monosomiese en een disomiese addisieplant is geïdentifiseer. Een AFLP- en twee RAPD-merkers is geïdentifiseer vir die Thinopyrum chromosoom(e) teenwoordig in die monosomiese addisieplant, terwyl drie AFLP- en drie RAPDmerkers geïdentifiseer is vir die disomiese addisieplant. 'n Poging is ook gemaak om diverse addisieplante te identifiseer met 'n volledige koroggenoom plus 'n addisionele Thinopyrum chromosoom. Plante met 2n = 43 / 44 is geïdentifiseer en gekarakteriseer met molekulêre merkers (AFLP en RAPD). Tros-analise is gebruik om die vermoedelik monosomiese of disomiese addisieplante te groepeer volgens die spesifieke Thinopyrum chromosome wat hulle behou het. Sewentien AFLP- en RAPD-merkers is gebruik om die 24 vermoedelike addisieplante in 6 groepe met verskillende Thinopyrum chromosome te groepeer. Alhoewel dit voorkom of die verskillende plante in 'n groep dieselfde addisionele Thinopyrum chromosoom het, is dit nie noodwendig die geval nie aangesien die interpretasie van die merkers bemoeilik word deur die heterogeniteit tussen die plante wat betref die agtergrond korogchromosome wat hulle besit. Dit is ook moontlik dat chromosoom herrangskikkings plaasgevind het gedurende die terugkruisings, wat die data verder kan bemoeilik. Nietemin, dit is nou moontlik om disomiese addisies te selekteer uit elke groep wat moontlik verskillende Thinopyrum chromosome bevat. Die data kan ook gebruik word om in die toekoms verdere addisieplante te identifiseer wat die oorblywende Thinopyrum chromosome bevat.

Thesis (MSc)--Stellenbosch University, 2004. / ENGLISH ABSTRACT: Much has been written about the effects of soil salinity on plant growth. Its devastating effects have already been reported 2000 years BC. In the 21· century an alarming 80 million hectares of cultivated land area are affected by salt (Munns, 2002a) and represent a growing threat to agriculture. Salt tolerance is a complex trait moderately expressed in only a few plant genotypes (Ruiz, 2001). An attempt to transfer salt tolerance genes from the wild grass, Thinopyrum distichum, to triticale and éommon wheat was initiated by Marais and Marais (2003). A study of Th. distichum x rye hybrids enabled the authors to identify chromosomes 2Jld , 3Jld , 4Jld and SJld as being involved in the determination of salt tolerance. Indirect (yet unconfirmed) evidence suggested that 7Jld might also have a role. A programme aiming to transfer regions of the critical chromosomes to homoeologous triticale chromosomes, which relies heavily on the use of molecular markers, was launched. While an RFLP marker is available for each of the Thinopyrum chromosomes, these are not suited for the screening of large numbers of segregates. This study therefore represents an attempt to convert the RFLP markers into less time consuming and cost-effective SCAR markers. The published DNA sequences of the RFLP probes in question were used as templates to design PCR primers. The PCR reactions were optimised using DNA of Th. distichum, rye and their FI hybrid. When Thinopyrum specific amplification products were obtained, the primers were also tested on a panel of genotypes with and without the target chromosomes. Seemingly polymorphic bands were confirmed by Southern blotting and hybridisation with the corresponding RFLP probes. The primers were also tested on a panel of genotypes that included 'Rex' triticale to ensure that they would also detect a difference in a triticale genetic background during transfer. Polymorphic bands were then isolated and sequenced to further refine the markers. In certain eases, sequences of the same fragment amplified in triticale ('Rex') and Thinopyrum were aligned in an attempt to design more specific markers. Using this approach, it was possible to develop chromosome specific SCARs for Thinopyrum chromosomes 3Jld and 7J2 d . Three and one set(s) of PCR markers, respectively, have been developed and can be used to unequivocally detect the Thinopyrum chromosomes involved in salt tolerance against a triticale background. A SCAR marker was also found for chromosome 6J. Thus, an attempt was made to convert thirteen RFLP probes to SCAR markers. Only three were successfully converted. The main reason for the low success rate is the syntenic relationships between the genomes of the different cereals that made it an arduous- task to find discriminating primer sets. Based on the results obtained, an adapted procedure is suggested for future attempts to develop chromosome specific markers utilizing published sequence information that was obtained for a different species. / AFRIKAANSE OPSOMMING: Baie is al geskryf oor die uitwerking van grond versouting op plantproduksie. Die vernietigende gevolge van versouting is alreeds 2000 jaar VC gerapporteer. In die 21* eeu is 'n geraamde 80 miljoen hektaar (Munns, 2002a) bewerkte land-area sout-geaffekteerd. Die ontstellende verwikkelinge verteenwoordig 'n groeiende bedreiging vir die landbou. Soutverdraagsaamheid is 'n komplekse kenmerk en slegs enkele plantgenotipes met matige verdraagsaamheid kon nog ontwikkel word (Ruiz, 2001). 'n Poging om soutverdraagsaamheidsgene vanaf die wilde gras, Thinopyrum distichum, na triticale en gewone koring oor te dra, is deur Marais en Marais (2003) geïnisieer. 'n Studie van Th. distichum x rog hibriede het die skrywers in staat gestelom chromosome (2Jld, 3Jld, 4Jld en SJld) wat bydra to soutverdraagsaamheid te identifiseer. Indirekte (maar onbevestigde) aanduidings is gevind dat 7J1dook' n rol mag speel. 'n Program is daarna geloods om segmente van chromosome na homoeoloë triticale chromosome oor te dra, 'n onderneming wat swaar steun op die gebruik van molekulêre merkers. Alhoewel daar'n RFLP merker beskikbaar is vir elk van die Thinopyrum chromosome, is hierdie merkers nie geskik vir die sifting van groot getalle segregate nie. Hierdie studie verteenwoordig 'n poging om die RFLP merkers om te skakel na 'n minder tydrowende en meer koste-effektiewe SCAR merkers. Die gepubliseerde DNS-volgordes van die betrokke RFLP peilers is as templaat gebruik om PKR inleiers te ontwerp. Die PKR reaksies is geoptimiseer deur gebruik te maak van DNS van Th. distichum. rog en hulle FI hibried. In gevalle waar Thinopyrum spesifieke amplifikasie produkte verkry is, is die inleiers ook getoets op 'n paneel van genotipes met en sonder die teikenchromosoom. Skynbare polimorfiese bande is bevestig deur 'n 'Southern' klad te maak en te hibridiseer met die tersaaklike RFLP peiler. Die inleiers is ook getoets op 'n paneel van genotipes waarby 'Rex' triticale ingesluit was om te verseker dat dit ook verskille in 'n triticale genetiese agtergrond opspoor (nodig tydens oordrag). Polimorfiese bande is verder verfyn. Dit is geïsoleer en die DNS-volgorde daarvan is bepaal. Tn sekere gevalle is ooreenstemmende fragmente geamplifiseer in triticale ('Rex') en Thinopyrum. Die volgordes is dan bepaal en met mekaar vergelyk in 'n poging om meer spesifieke merkers te ontwerp. Met die gebruik van hierdie benadering was dit moontlik om chromosoom-spesifieke SCAR-merkers vir die Thinopyrum chromosome 3Jld en 7J2d te ontwikkel. Drie en een stel(le) PKR merkers is onderskeidelik ontwikkel en kan gebruik word om ondubbelsinnig te bepaal of die betrokke Thinopyrum chromosoom segregeer in 'n triticale kruising. 'n SCAR merker is ook gevind vir chromosoom 6J. Dus, daar is probeer om dertien RFLP peilers na SCAR merkers om te skakel. Slegs drie van die pogings was suksesvol. Die hoofrede vir die lae sukseskoers is die hoë graad van sintenie tussen die genome van die verskillende grane wat dit 'n moeilike taak gemaak het om diskriminerende inleierstelle te ontwerp. Op grond van die resultate word 'n ietwat gewysigde prosedure vir die toekomstige pogings om chromosoom-spesifieke merkers te ontwerp met gebruik van gepubliseerde volgorde inligting vanaf' n ander spesie, voorgestel.

[Truncated abstract] Dryland salinity is a major environmental challenge facing agriculture in Australia. One option to manage dryland salinity is the use of perennial forages that increase water use of agricultural systems. However, the current array of perennial forages is limited. Forage species that satisfy the range of climatic and edaphic environments, and production systems, in southern Australia are needed (Chapter 1). In particular, low rainfall regions lack options other than lucerne (Medicago sativa L.) (Chapter 1). The Dorycnium genus (canary clovers) contains perennial species that might be useful forage plants for southern Australia. Dorycnium are sub-shrubs and their plant form differs from current perennial forages (Chapter 1). The aim of this project was to investigate some of the agronomic traits of several species of the genus Dorycnium to explore where they might be used in Australia and how they might be integrated into agricultural systems for management of dryland salinity. First, two desktop investigations assessed the potential adaptation and role of Dorycnium species in southern Australia: a review of the current literature on the agronomic characteristics of Dorycnium (Chapter 2) and an eco-geographical analysis to explore the ecology of Dorycnium species (Chapter 3). The agronomy of Dorycnium has been previously researched mainly in New Zealand, and although this provides some indications on where and how Dorycnium might be best used in Australia, this still requires testing in Australia. In particular, the aluminium tolerance of Dorycnium species indicates that they may be more suitable for acid soils than lucerne. Little ecological data was obtained for germplasm and herbarium collection sites of Dorycnium species. Climate comparisons between the native distribution of Dorycnium species in the Mediterranean basin and Australia, using spatial aridity data and CLIMEX climate match modelling, revealed that D. hirsutum and D. rectum might be suitably adapted to the temperate pasture regions of southern Australia. Suitable germplasm of D. pentaphyllum may also exist, but subsequent investigations in this project focussed on D. hirsutum and D. rectum.

Soil salinity and the arid climate in Libya are major constraints in agriculture and predominantly in foodstuff production which are limiting wheat production and yield. The effect of pre-sowing seed treatments with 50 mM of KCl, NaCl, CaCl2, and distilled water as hydropriming on germination and early seedling growth in two wheat (Triticum aestivum L.) cultivars S-24 (tolerant) and Slambo (untested before) under 0, 100, 200 and 300 mM NaCl concentrations was examined. CaCl2 was the only priming treatment that significantly improved the germination percentage, germination rate, and mean germination time in both cultivars under almost all NaCl concentrations. Thus, priming with CaCl2 was selected for further experiments. In the greenhouse, seeds primed with 50 mM of CaCl2 also improved the emergence percentage, emergence rate, shoot and root length, and fresh and dry weight of shoots and roots of both cultivars under all NaCl concentration except at 300 mM where the emergence was completely inhibited. The response of wheat cultivars to three compost treatments including cow manure compost (CC), greenwaste compost (GC) and 50:50 mixture (mix) between them and sand at percentage inclusions of 10 and 30 % by weight under 0, 100, 200, and 300 mM of NaCl was also investigated. Among all compost treatments, 30% GC and 30% mix were the best treatment and improved almost all growth parameters under salt stress, and 30% GC was also the only treatment that had any emergence at 300 mM NaCl. 30% GC and 30% mix were selected for further experiments. The effect of the combination of the selected priming agent (CaCl2) and the best two compost treatments (30% GC and 30% mix) on the emergence and early seedling growth of both cultivars was tested. The results showed that all the treatments enhanced plant growth parameters including seedling ion uptake in both cultivars, with preference to primed seeds sown in 30% GC. The treatments had the following order of the performance of both cultivars under salt stress. Primed seeds sown in 30% GC > unprimed seeds sown in 30% GC > primed seeds sown in 30% mix > unprimed seeds sown in 30% mix. This enhancement is possibly due to the provision of Ca2+ and / or the improvement in the availability of water as both of them were improved by the application of priming and compost.

Two problems hampering efforts to produce salt-tolerant plants through constitutive expression of transgenes include: 1. Spatial control. Particular cell-types must respond specifically to salt stress to minimise the amount of Na⁺ delivered to the shoot; and, 2. Temporal control. Transgenes are typically expressed in plants at similar levels through time, irrespective of the stress encountered by the plant, which may exacerbate pleiotropic effects and means that, particularly in low-stress conditions, costly and/or detrimental metabolic processes may be active, thus reducing yield. To address these issues, Gateway® destination vector constructs were developed combining the GAL4 UAS (upstream activating sequence) with the ethanol-inducible gene expression system to drive inducible cell-specific expression of Na⁺ transporter transgenes (or to silence salt transporter transgenes inducibly and cell-specifically). Rice (Oryza sativa L. cv. Nipponbare) GAL4-GFP enhancer trap lines (Johnson et al., 2005: Plant J. 41, 779-789) that express GAL4 and GFP specifically in either the root epidermis or xylem parenchyma (and therefore ‘trap’ cell-type specific enhancer elements) were transformed with this GAL4 UAS – ethanol switch construct, thereby allowing both spatial and temporal control of transgenes. In preliminary experiments, the expression system successfully limited the expression of RFP to specific cell-types after induction with ethanol. Other genes expressed using this system include PpENA1, a Na⁺-extruding ATPase from the moss, Physcomitrella patens, and AtHKT1;1, a Na ⁺ transporter from Arabidopsis thaliana. The two enhancer trap rice lines were also transformed with the GAL4 UAS driving stable expression of AtHKT1;1 and PpENA1 specifically in root epidermal or xylem parenchyma cells. Expression of AtHKT1;1 in root epidermal cells reduced Na⁺ accumulation in the shoots, while expression in the root xylem parenchyma appeared to have little effect on shoot Na⁺ accumulation. Using cryo-scanning electron microscopy (SEM) X-ray microanalysis, the outer cells of the roots of the line expressing AtHKT1;1 in the epidermal cells were found to accumulate higher levels of Na⁺ than the parental enhancer trap line. Additionally, this line had decreased unidirectional ²²Na⁺ influx. Similar results were observed for plants expressing AtHKT1;1 driven by the CaMV 35S / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1325289 / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2008

Two problems hampering efforts to produce salt-tolerant plants through constitutive expression of transgenes include: 1. Spatial control. Particular cell-types must respond specifically to salt stress to minimise the amount of Na⁺ delivered to the shoot; and, 2. Temporal control. Transgenes are typically expressed in plants at similar levels through time, irrespective of the stress encountered by the plant, which may exacerbate pleiotropic effects and means that, particularly in low-stress conditions, costly and/or detrimental metabolic processes may be active, thus reducing yield. To address these issues, Gateway® destination vector constructs were developed combining the GAL4 UAS (upstream activating sequence) with the ethanol-inducible gene expression system to drive inducible cell-specific expression of Na⁺ transporter transgenes (or to silence salt transporter transgenes inducibly and cell-specifically). Rice (Oryza sativa L. cv. Nipponbare) GAL4-GFP enhancer trap lines (Johnson et al., 2005: Plant J. 41, 779-789) that express GAL4 and GFP specifically in either the root epidermis or xylem parenchyma (and therefore ‘trap’ cell-type specific enhancer elements) were transformed with this GAL4 UAS – ethanol switch construct, thereby allowing both spatial and temporal control of transgenes. In preliminary experiments, the expression system successfully limited the expression of RFP to specific cell-types after induction with ethanol. Other genes expressed using this system include PpENA1, a Na⁺-extruding ATPase from the moss, Physcomitrella patens, and AtHKT1;1, a Na ⁺ transporter from Arabidopsis thaliana. The two enhancer trap rice lines were also transformed with the GAL4 UAS driving stable expression of AtHKT1;1 and PpENA1 specifically in root epidermal or xylem parenchyma cells. Expression of AtHKT1;1 in root epidermal cells reduced Na⁺ accumulation in the shoots, while expression in the root xylem parenchyma appeared to have little effect on shoot Na⁺ accumulation. Using cryo-scanning electron microscopy (SEM) X-ray microanalysis, the outer cells of the roots of the line expressing AtHKT1;1 in the epidermal cells were found to accumulate higher levels of Na⁺ than the parental enhancer trap line. Additionally, this line had decreased unidirectional ²²Na⁺ influx. Similar results were observed for plants expressing AtHKT1;1 driven by the CaMV 35S / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1325289 / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2008