Although plant growth-promoting rhizobacteria (PGPR) have shown tremendous potential to be used as inoculants for many agricultural crops, they may not survive severe environmental conditions in the field which could limit their large scale applications. Endophytic bacteria, which can be recovered from inside plant tissues such as roots, stems and leaves, might overcome this limitation due to their unique ecological niche inside plant roots where they are sheltered from external environmental disturbances. Some of these bacterial endophytes have beneficial effects on their host plants and stimulate plant growth or reduce disease symptoms, apparently through mechanisms that are similar to those proposed for PGPR. The objective of this study was to assess a collection of endophytic bacteria for PGPR traits and potential use to enhance the rhizobial-legume symbiosis. Forty isolates obtained from the roots of various plants were identified by fatty acid methyl ester (FAME) analysis, and 16S RNA gene sequencing analysis. The majority (i.e., 75%) were identified as Pseudomonas species. Many of these isolates were able to solubilize phosphate, produce indole-3-acetic acid (IAA), produce aminocyclopropane-1-carboxylic acid (ACC) deaminase, synthesize siderophores and show antagonistic activities against several soil-borne plant pathogenic fungi under in vitro conditions. Selected isolates were further evaluated for the ability to enhance plant growth and nodulation of alfalfa when co-inoculated with Sinorhizobium meliloti under growth chamber conditions using growth pouch and potted soil assays. Results revealed that P. putida strain EB EE 4-25, P. syringae strain EB XDE 4-48, and P. fluorescens strain EB EE 2-23 significantly increased shoot length, root length, enhanced nodulation and increased lateral root formation of alfalfa plants in growth pouch and potted soil assays when co-inoculated with S. meliloti strain P102 compared to plants inoculated with S. meliloti strain P102 alone. Results also suggested that expression of one or more of the mechanisms, such as solubilization of phosphate, production of IAA, production of siderophores, and ACC deaminase production might have played a role in the enhancement of the alfalfa- Sinorhizobium symbiosis. These results suggest that some endophytic bacterial strains may be useful as biofertilizers and/or biocontrol agents in sustainable agricultural practices.

Although plant growth-promoting rhizobacteria (PGPR) have shown tremendous potential to be used as inoculants for many agricultural crops, they may not survive severe environmental conditions in the field which could limit their large scale applications. Endophytic bacteria, which can be recovered from inside plant tissues such as roots, stems and leaves, might overcome this limitation due to their unique ecological niche inside plant roots where they are sheltered from external environmental disturbances. Some of these bacterial endophytes have beneficial effects on their host plants and stimulate plant growth or reduce disease symptoms, apparently through mechanisms that are similar to those proposed for PGPR. The objective of this study was to assess a collection of endophytic bacteria for PGPR traits and potential use to enhance the rhizobial-legume symbiosis. Forty isolates obtained from the roots of various plants were identified by fatty acid methyl ester (FAME) analysis, and 16S RNA gene sequencing analysis. The majority (i.e., 75%) were identified as Pseudomonas species. Many of these isolates were able to solubilize phosphate, produce indole-3-acetic acid (IAA), produce aminocyclopropane-1-carboxylic acid (ACC) deaminase, synthesize siderophores and show antagonistic activities against several soil-borne plant pathogenic fungi under in vitro conditions. Selected isolates were further evaluated for the ability to enhance plant growth and nodulation of alfalfa when co-inoculated with Sinorhizobium meliloti under growth chamber conditions using growth pouch and potted soil assays. Results revealed that P. putida strain EB EE 4-25, P. syringae strain EB XDE 4-48, and P. fluorescens strain EB EE 2-23 significantly increased shoot length, root length, enhanced nodulation and increased lateral root formation of alfalfa plants in growth pouch and potted soil assays when co-inoculated with S. meliloti strain P102 compared to plants inoculated with S. meliloti strain P102 alone. Results also suggested that expression of one or more of the mechanisms, such as solubilization of phosphate, production of IAA, production of siderophores, and ACC deaminase production might have played a role in the enhancement of the alfalfa- Sinorhizobium symbiosis. These results suggest that some endophytic bacterial strains may be useful as biofertilizers and/or biocontrol agents in sustainable agricultural practices.

Magister Scientiae - MSc (Biotechnology) / Endophytic bacteria are known to have an endosymbiotic relationship with plants and provide them with many beneficial properties. These bacteria stimulate plant hormones, provide protection from pathogens and increase nutrient availability in the environment. In this study some of these potential growth factors were tested. Endophytic bacteria have the potential to be of great value for the increase of crop production. They offer a variety of processes that aid in plant growth promotion in an ecofriendly manner. The use of endophytic bacteria provides a cheaper and cleaner approach compared to industrial made fertilizers. They also have potential uses in bioremediation to clean the environment polluted by industrial processes. Endophytes were isolated and showed significant growth improvement. Each isolate displayed different morphologies. Isolates were tested for classical growth promotion mechanisms such as the ability to solubilize phosphate, Indole-3-acetic acid and siderophore production. Inductively Coupled Plasma Optical Emission Spectrometry was performed to measure the effect of the isolates on the plants nutrient profile. The isolates were then tested again while the plants were under heavy metal stress to determine if they were still capable of growth promotion. The plants were then assayed for cell death using Evans blue and biomass was measured to determine the effect of vanadium stress. Inductively Coupled Plasma Optical Emission Spectrometry was performed again to assess the change in nutrient profile while under vanadium stress. / 2021-08-31

Expanding global populations, unequal food distribution and disease pressure suggest food poverty is increasing. Consequently, much attention is focussed on alternative natural methods in which to increase agricultural yield. Previously, it was observed that Trichoderma hamatum strain GD12 and its respective N-acetyl-β-D-Glucosamine mutant ∆Thnag:hph promoted plant biomass and fitness that, as a result, may provide a credible natural alternative to synthetic fertilisers. However, on a molecular level, the manner in which this is achieved has not been fully elucidated. In this thesis, I report the biofertiliser effect of GD12 and mutant ∆Thnag::hph once applied to autoclaved peat microcosms as sole applications. Furthermore, I demonstrate the biocontrol ability of GD12 when co-inoculated with Sclerotinia sclerotiorum or Rhizoctonia solani and reveal, that once mycelium co-inoculation has occurred, GD12 increase plant biomass and provide protection; whilst ∆Thnag::hph does not. Consequently, I challenged the biocontrol effects of Trichoderma metabolite extract where I validate that both Trichoderma wild type GD12 and mutant ∆Thnag::hph are incapable of suppressing pathogen growth. Subsequently, I characterised the up-regulated signatures associated with GD12 and ∆Thnag::hph using LC-MS techniques where unique compounds were discovered from each strain of Trichoderma. In conclusion, I provide evidence that N-acetyl-β-D-Glucosamine mutation bring about metabolomic changes that affect the fungal secretome which, in turn, alters plant phenotype, fitness and germination. Furthermore, I have shown that these effects are species specific and depend upon pathogen, plant and fungal properties. However, further investigations are needed to fully elucidate the compound(s) responsible for biocontrol and biofertilisation; especially plant-specific effects that take place as a consequence of fungal activity.

Thesis (MSc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: In order to test the effect of different plant growth-promoting strategies on Triticum aestivum L. (wheat), we investigated the ability of biochar and a grain-associated soil yeast, to improve the growth of this crop. Our first goal was to study the effect of biochar amendments to sandy soil on the growth and nutrition of wheat in the presence of mycorrhizal fungi. This was accomplished by amending soil with 0%, 1%, 2.5%, 5% and 10% (w/w) biochar and cultivating wheat plants in these soil-biochar mixtures. After harvesting, plant growth and mycorrhizal colonization of roots were measured. In addition, we studied the nutritional physiology of these plants with regards to nitrogen (N), phosphorous (P) and potassium (K) concentrations, as well as the growth efficiencies and uptake rates of these nutrients. We found that wheat growth was improved by biochar amendments to soil, probably as a result of elevated K levels in the plant tissues supplied by the biochar amendments. The second goal of this study was to obtain a soil yeast from the rhizosphere of another monocot in the family Poaceae, i.e. Themeda triandra Forssk. (red grass), and then evaluate this isolate for its ability to improve wheat performance. Three different Cryptococcus species were isolated from the rhizosphere of wild grass, i.e. Cryptococcus zeae, Cryptococcus luteolus and Cryptococcus rajasthanensis. Since C. zeae was previously isolated from maize, an isolate representing this species was selected to be used in further experimentation. With the ultimate goal of testing the ability of this yeast to improve wheat growth, its effect on wheat germination was investigated and compared to that of two other soil yeasts, i.e. Cryptococcus podzolicus CAB 978 and Rhodotorula mucilaginosa CAB 826. These three yeasts were subsequently tested for their ability to improve wheat growth in pot cultures in a greenhouse. After one and two months of growth, the culturable yeasts present in the rhizosphere and bulk soil were enumerated. The effects of these yeasts were elucidated by measuring wheat growth in terms of dry weight, as well as root and shoot relative growth rates (RGR). Changes in wheat nutrition were evaluated by determining the concentrations, growth efficiencies and uptake rates for P, K, zinc (Zn) and iron (Fe). During this study, it was found that only C. zeae CAB 1119 and C. podzolicus CAB 978 were able to enhance seed germination. Similarly, it was shown that C. zeae CAB 1119 was able to improve wheat growth during the first and second month of cultivation, whereas C. podzolicus CAB 978 only improved growth during the first month, and R. mucilaginosa CAB 826 had no effect on growth. This improved growth could be attributed to C. zeae CAB 1119 improving the P, K, Zn and Fe growth efficiency of wheat, which positively influenced the root and shoot RGR, and subsequently wheat growth. Our final goal was to test whether C. zeae CAB 1119 could affect wheat growth and nutrition when cultivated in sandy soil, which contained natural microbial consortia and 10% (w/w) biochar. Plants treated with viable or autoclaved cells of C. zeae CAB 1119, were subsequently cultivated in soil only or soil amended with biochar. After one month, plants were harvested and growth was measured with regards to dry weight, root RGR and shoot RGR. In addition, the concentrations of P, K, Zn and Fe were analyzed for these plants, where after the growth efficiencies and uptake rates were calculated for these four nutrients. Results indicated that plants growing in soil amended with biochar, and treated with viable C. zeae CAB 1119, showed the best growth. The increased root and shoot RGR witnessed in these plants was probably due to increased concentrations of P and K in the plants. This study opens new avenues of research with regards to the bio-fertilizers of wheat. / AFRIKAANSE OPSOMMING: Die uiteindelike doel van die studie was om die effek van verskillende plantgroei bevorderende metodes op die groei van Triticum aestivum L. (koring) te ondersoek. Dus het ons die vermoë van houtskool en ‘n graan-geassosieerde grondgis getoets om die groei van dié plant te bevorder. Die eerste doel van die studie was om die effek van houtskool toedienings tot sanderige grond te evalueer. Dit is bewerkstellig deur 0%, 1%, 2.5%, 5% en 10% (w/w) van die houtskool by die sand toe te voeg en koring in die houtskool-sand mengsels te kweek. Na die verlangde groei tydperk is die koring geoes en die mikorrizale kolonisasie op en in die koring wortels bepaal. Gedurende hierdie studie is die effek van bogenoemde toedienings op die fisiologie van die plante ondersoek deur die konsentrasies, opname tempo’s, en groei ekonomie van die plante vir stikstof (N), fosfaat (P) en kalium (K) te bepaal. Ons het gevind dat die groei van koring deur die toediening van houtskool bevorder is en dit blyk dat dié effek weens die teenwoordigheid van hoë K vlakke in die plantweefsel is. Die tweede doel van ons studie was om ‘n gis vanuit die risosfeer van ‘n monokotiel wat aan die familie Poacea behoort, naamlik Themeda triandra Forssk. (rooigras) te isoleer. Die vermoë van die isolaat om die groei van koring te bevorder was daarna getoets. Drie verskillende Cryptococcus spesies was vanuit die risosfeer van rooigras geïsoleer, nl. Cryptococcus zeae, Cryptococcus luteolus en Cryptococcus rajasthanensis. Omdat C. zeae in ‘n vorige studie vanaf mielies geisoleer was, is ‘n isolaat van hierdie spesie gebruik in verdere eksperimente. Met die doel om te bepaal of dié gisspesie koringgroei kan bevorder, was die effek van C. zeae op die ontkieming van koring bestudeer en vergelyk met dié van twee ander grond giste, nl. Cryptococcus podzolicus CAB 978 en Rhodotorula mucilaginosa CAB 826. Hierdie drie giste is ook ondersoek om die groei van koring in ‘n glashuis te bevorder. Na een en twee maande se groei was die getalle van giste teenwoordig in die risosfeer en grond verder weg van die wortels bepaal. Die effek van dié giste op die groei van koring is bepaal in terme van droë gewig asook die relatiewe wortel en halm groei tempos. Veranderinge in die nutrient status van koring is ondersoek deur die konsentrasies, groei-ekonomie en tempo van opname vir P, K, sink (Zn) en yster (Fe) te bepaal. Ons het gedurende dié studie gevind dat C. zeae CAB 1119 en C. podzolicus CAB 978 die ontkieming van koring kon verbeter. Ons het ook gevind dat C. zeae CAB 1119 die groei van koring gedurende die eerste en tweede maand van groei kon bevorder, terwyl C. podzolicus CAB 978 dit net gedurende die eerste maand kon vermag en R. mucilaginosa CAB 826 geen effek gehad het nie. Die verbeterde groei kon aan C. zeae CAB 1119, wat die P, K, Zn en Fe groei effektiwiteit van die plante verbeter het, toegeskryf word. Die verbetering van groei effektiwiteit het ‘n positiewe invloed op die relatiewe groeisnelheid van die wortels en halms gehad, en dus op koringgroei. Die laaste doel van die studie was om te bepaal of C. zeae CAB 1119 die groei van koring kon bevorder wanneer die koring in sand wat natuurlike mikrobiese populasies bevat en met houtskool aangevul is, gekweek word. Plante is met lewensvatbare of nielewensvatbare selle van C. zeae CAB 1119 behandel en gekweek in sanderige grond, en/of grond waarby 10% (w/w) houtskool toegevoeg is. Die plante is na een maand geoes en die groei bepaal in terme van droë massa en die relatiewe wortel en halm groei tempos. Die konsentrasies van P, K, Zn en Fe in die plante, asook die fisiologie van die plante, nl. groei ekonomie en tempo van opname, met betrekking tot P, K, Zn en Fe is bepaal, Ons het gevind dat plante wat in die houtskool-grond mengsel gekweek is en met lewensvatbare selle van C. zeae CAB 1119 behandel is die beste groei getoon het. Die verbeterde relatiewe groei tempos van die wortels en halms was mees waarskynlik die gevolg van verhoogde P en K konsentrasies in die plante. Hierdie studie toon nuwe resultate in verband met die gebruik van biologiese alternatiewes tot kunsmis.

Trichoderma species are soil-borne filamentous fungi widely utilized for their many plant health benefits, such as conferring improved growth, disease resistance and abiotic stress tolerance to their hosts. Many Trichoderma species are able to produce the auxin phytohormone indole-3-acetic acid (IAA), and its production has been suggested to promote root growth. Here we show that the production of IAA is strain dependent and diverse external stimuli are associated with its production. In in vitro assays, Arabidopsis primary root length was negatively affected by the interaction with some Trichoderma strains. In soil experiments, a continuum effect on plant growth was shown and this was also strain dependent. In plate assays, some strains of Trichoderma spp. inhibited the expression of the auxin reporter gene DR5 in Arabidopsis primary roots but not secondary roots. When Trichoderma spp. and A. thaliana were physically separated, enhancement of both shoot and root biomass, increased root production and chlorophyll content were observed, which strongly suggested that volatile production by the fungus influenced the parameters analyzed. Trichoderma strains T. virens Gv29.8, T. atroviride IMI206040, T. sp. "atroviride B" LU132, and T. asperellum LU1370 were demonstrated to promote plant growth through volatile production. However, contrasting differences were observed with LU1370 which had a negative effect on plant growth in soil but a positive effect in plate assays. Altogether our results suggest that the mechanisms and molecules involved in plant growth promotion by Trichoderma spp. are multivariable and are affected by the environmental conditions.

Plants naturally host complex microbial communities in which the plant and the symbiotic partners act as an integrated metaorganism. These communities include beneficial (i.e. plant growth promoting, PGP) microorganisms which provide fundamental ecological services able to enhance host plant fitness and stress tolerance. PGP microorganisms represent a potential bioresource for agricultural applications, especially for desert farming under the harsh environmental conditions occurring in hot/arid regions (i.e. drought and salinity). In this context, understanding the ecological aspects of the associated microorganisms is crucial to take advantage of their ecological services. Here, hot/desert ecosystems were selected and two contrasting plant categories were used as models: (i) wild plants (i.e. speargrasses) growing in hot-desert sand dunes and (ii) the main crop cultivated in desert ecosystems, the date palm. By using highthroughput DNA sequencing and microscopy, the ecology and functionality of the microbial communities associated with these plants were characterized. Additionally, the PGP services of bacteria isolated from date palm were explored. I found that the harsh conditions of the desert strongly affect the selection and assembly of microbial communities associated with three different speargrass species, determining a plant species-independent core microbiome always present among the three plant species and carrying important PGP traits. On the contrary, in agroecosystems where desert farming practices are used, the plant species, i.e. date palm exerts a stronger selective pressure than the environmental and edaphic factors favoring the recruitment of conserved microbial assemblages, independent of the differences in the soil and environmental conditions among the studied oases. Such selective pressure also favors the recruitment of conserved PGP microorganisms (i.e. Pseudomonas sp. bacterial strains) able to protect their host from salinity stress through the induction of root architectural changes regulated by the modification of the root system auxin homeostasis. Overall, we found that deserts are unique ecosystems that challenge the paradigm of microbial community assembly, as it was defined from studies in non-arid ecosystems. The understanding of the ecological features regulating the ecological properties of such unique microbial community assembly will be a key-step to improve the chances of successful application of ‘PGP microorganisms’ in arid agroecosystems.

<p><i>Paenibacillus polymyxa</i> belongs to the group of plant growth promoting rhizobacteria (PGPR). Activities associated with <i>P. polymyxa</i>-treatment of plants in earlier experiments include, e.g., nitrogen fixation, soil phosphorus solubilization, production of antibiotics, auxin, chitinase, and hydrolytic enzymes, as well as promotion of increased soil porosity. My thesis work showed that, in stationary phase, <i>P. polymyxa</i> released the plant hormone cytokinin isopentenyladenine, in concentrations of about 1.5 nM.</p><p>In a gnotobiotic system with <i>Arabidopsis thaliana</i> as a model plant, it was shown that <i>P. polymyxa</i>-inoculation protects plants; challenge by either the pathogen <i>Erwinia carotovora</i> (biotic stress) or induction of drought (abiotic stress) showed that pre-inoculated plants were significantly more resistant than control plants. By RNA-differential display on RNA from <i>P. polymyxa</i>-treated or control plants, changes in gene expression were tested. One mRNA, encoding ERD15 (drought stress-responsive gene) showed a strong inoculation-dependent increase in abundance. In addition, several biotic stress-related genes were also activated by <i>P. polymyxa</i>. </p><p>Antagonism towards the fungal pathogens <i>Phytophthora palmivora</i> and <i>Pythium aphanidermatum</i> was studied. <i>P. polymyxa</i> counteracted the colonization of zoospores of both oomycetes on <i>A. thaliana</i> roots, and survival rates of plants treated with <i>P. polymyxa</i> were much higher when challenged by <i>P. aphanidermatum</i>. </p><p>Using a green fluorescent protein-tagged isolate of <i>P. polymyxa</i>, colonization of <i>A. thaliana</i> roots was investigated. Two main conclusions can be drawn. Firstly, the bacterium enters the root tissue (but not leaves) and is abundantly present in intercellular spaces. Secondly, the root becomes severely damaged, indicating that – under some conditions – <i>P. polymyxa</i> is a "deleterious bacterium", and in others it promotes growth. Based on work presented in my thesis, I argue that a balance between the activities of a PGPR, the genetic background and physiological state of a plant, and the environmental conditions employed in test systems, ultimately determines the resulting effect. </p>

Paenibacillus polymyxa belongs to the group of plant growth promoting rhizobacteria (PGPR). Activities associated with P. polymyxa-treatment of plants in earlier experiments include, e.g., nitrogen fixation, soil phosphorus solubilization, production of antibiotics, auxin, chitinase, and hydrolytic enzymes, as well as promotion of increased soil porosity. My thesis work showed that, in stationary phase, P. polymyxa released the plant hormone cytokinin isopentenyladenine, in concentrations of about 1.5 nM. In a gnotobiotic system with Arabidopsis thaliana as a model plant, it was shown that P. polymyxa-inoculation protects plants; challenge by either the pathogen Erwinia carotovora (biotic stress) or induction of drought (abiotic stress) showed that pre-inoculated plants were significantly more resistant than control plants. By RNA-differential display on RNA from P. polymyxa-treated or control plants, changes in gene expression were tested. One mRNA, encoding ERD15 (drought stress-responsive gene) showed a strong inoculation-dependent increase in abundance. In addition, several biotic stress-related genes were also activated by P. polymyxa. Antagonism towards the fungal pathogens Phytophthora palmivora and Pythium aphanidermatum was studied. P. polymyxa counteracted the colonization of zoospores of both oomycetes on A. thaliana roots, and survival rates of plants treated with P. polymyxa were much higher when challenged by P. aphanidermatum. Using a green fluorescent protein-tagged isolate of P. polymyxa, colonization of A. thaliana roots was investigated. Two main conclusions can be drawn. Firstly, the bacterium enters the root tissue (but not leaves) and is abundantly present in intercellular spaces. Secondly, the root becomes severely damaged, indicating that – under some conditions – P. polymyxa is a "deleterious bacterium", and in others it promotes growth. Based on work presented in my thesis, I argue that a balance between the activities of a PGPR, the genetic background and physiological state of a plant, and the environmental conditions employed in test systems, ultimately determines the resulting effect.

Thesis (MSc (Plant Biotechnology))--University of Stellenboscg, 2010. / Includes bibliography. / Title page: Dept. of Genetics, Faculty of Natural Sciences. / ENGLISH ABSTRACT: Novel plant growth regulating substances (PGRs) are emerging as a useful tool to investigate important growth traits in plants. This study reports on growth promotion pathways leading to enhanced biomass accumulation in two PGRs sharing a common α, β-unsaturated furanone moiety. Growth promotion by GR24, a synthetic strigolactone, and an aqueous smoke solution (including the active compound, KAR1) in physiologically normal seedlings was characterized by enhanced biomass accumulation and higher seedling vigour. Root architecture (lateral root number and root length) and shoot size (fresh and dry shoot weight and leaf area) were also dramatically improved following GR24 and smoke/KAR1 treatment. Despite these apparent similarities, parallel transcript and phytohormone profiling identified only a limited number of overlapping entities. Four common up-regulated and nineteen down-regulated mRNA transcripts were identified; whilst amongst the phytohormones that were analyzed, only ABA and JA levels were commonly increased between the treatments. This suggests that, whilst the phenotypic end response(s) was similar, it was attained via distinct pathways. The limited number of co-expressed transcripts between these treatments, as well as repressed biomass accumulation when combining GR24 and aqueous smoke in a single treatment suggests, however, that a certain degree of cross-talk in either signal perception/transduction and/or biomass regulation could not be ruled out. In light of the structural similarity between the strigolactone and KAR1 molecules and the degree of redundancy between these treatments, it is possible that these two molecules might share a common receptor/perception pathway. Two silencing vectors were constructed, specifically aimed at silencing Nicotiana benthamiana genes MAX4 and MAX2 which are known to function in the strigolactone biosynthesis pathway and signal transduction pathway, respectively. Transgenes designed to express single- or double-stranded-self- complementary hairpin RNA have a post translational gene silencing effect. The pHELLSGATE2 plasmid a binary vector that incorporates GATEWAY cloning technology which makes use of λ-phage-based site specific recombination, rather than restriction endonucleases and ligation, was used to construct these gene silencing vectors. These constructs can in future be used to produce Nicotiana plants with impaired strigolactone production and perception abilities and may provide evidence as to whether the signaling cascade of KAR1 and strigolactone share a degree of crosstalk. / AFRIKAANSE OPSOMMING: Aanvraag na plantmateriaal is besig om toe te neem, hetsy vir gebruik as mens- en diervoeding of vir die produksie van biobrandstof. Om aan hierdie behoefte te voldoen, word verskeie pogings geloods wat fokus op die optimisering van plantproduksiestelsels. Om plantgroei te stimuleer/verbeter, is ’n ingewikkelde proses en is oor die algemeen moeilik om te begryp. Die produksie van plantbiomassa is nou gekoppel aan primêre metabolisme en enige verandering in hierdie biochemiese padweë kan lei tot ongewenste newe-effekte. Gevolglik word primêre metabolisme streng beheer deur reguleringsmeganismes. ’n Nuttige alternatief tot metaboliese wysiging is deur bio-aktiewe agente te karakteriseer op grond van die veranderinge aan plantgroei wat waargeneem word. Nuwe stowwe met biologiese aktiwiteite in plantontwikkeling word elke dag ontdek en speel ’n belangrike rol in die studie van plantgroei en -ontwikkeling. Hier word verslag gelewer van twee plantgroei-stimulerende stowwe wat albei lei tot die aktivering van verbeterde plantbiomassa-akkumulasie-padweë. Swaarder plantjies met ’n verhoogde oorlewingsvermoё is waargeneem in fisiologies normale saailinge wat met ’n sintetiese strigolaktoon (GR24) of met rookwater (met aktiewe bestanddeel, KAR1) behandel is. Behandeling met hierdie twee stowwe het gelei tot soortgelyke plantbiomassa-akkummulasie- vermoё. Hierdie twee stowwe (GR24 en KAR1) deel ’n ooreenstemmende molekulêre struktuur in die vorm van ’n α, β-onversadigde furanone-moieteit. Ten spyte van die groeiverbeteringsooreenkomste, gesien in saalinge behandel met GR24 en rook/KAR1, dui verskille in transkripsie- en hormoonprofiel op twee verskillende groeistimuleringspadweë. Saailinge wat gelyktydig behandel is met ’n kombinasie van die twee stowwe het egter ’n stremming in groei getoon in vergelyking met die kontroleplantjies. Dit is egter waargeneem dat daar wel ’n mate van oorvleueling in die aantal transkripte was tussen die drie behandelinge, wat daarop dui dat die groei-regulerende padweë nie in totale onafhanklikheid funksioneer nie, maar wel sekere stappe deel. Na aanleiding van die strukturele ooreenkomste tussen die strigolaktoon (GR24) en KAR1 molekules en die mate van molekulêre kommunikasieoorvleueling word gepostuleer dat hierdie twee molekules dalk aan dieselfde reseptormodule kan bind of stimuleer. Om hierdie rede is twee geendempingsvektors geskep wat daarop gemik is om twee gene, MAX2 en MAX4, in Nicotiana benthamiana uit te doof. Die MAX2 geenproduk is betrokke in die kommunikasie en waarneming van die strigolaktoon en die MAX4 geenproduk is betrokke by die vervaardiging van die hormoon. Oordraagbare geen-kostruksies wat daarop gemik is om enkel- en dubbelstring selfkomplimentêre haarnaald-RNS te vorm, besit die vermoë om getranskribeerde geenprodukte te vernietig. Die pHELLSGATE2 plasmied is ’n binêre vektor wat GATEWAY kloneringstegnologie gebruik, waar λ-faag gebaseerde setelspesifieke rekombinasie eerder as die tradisionele ligeringsreaksie gebruik word. Hierdie konstrukte kan gebruik word om transgeniese plantjies te skep waar die vermoë om strigolaktoon te maak of waar te neem, verloor of onderdruk is. Hierdie transgeniese plantjies kan gebruik word om te bepaal of die plantgroei-stimulerende vermoë van GR24 en rook/KAR1 wel dieselfde padweë gebruik.