Immersion System Research Articles

Novel Corrosion Inhibitor for Mild Steel in Simulated Concrete Pore Solution with Chloride Contamination

The corrosion resistance performance of mild steel in simulated concrete pore solution (pH >12) containing a) 0.5 M sodium Nitrite b) 0.2 M sodium citrate c) 0.4 M sodium benzoate inhibitors in 0.5 M NaCl solution contamination were evaluated by electrochemical studies. The observed inhibition efficiencies were compared with gravimetric method after 30 days of immersion, for inhibited and uninhibited system. The maximum inhibition efficiency of 0.5 M sodium nitrite +0.2 M sodium citrate +0.4 M sodium benzoate mixed inhibitor system was found to be 75%. The inhibition efficiencies for the mixed inhibitor system were compared with control and single inhibitor system. From these study it was found that the role of nitrite ion present in the mixed inhibitor system repassivated the mild steel that drastically reduce the corrosion rate even in presence of high chloride concentration.

Micropropagation of Embryogenic Callus of Oil Palm (Elaeis Guineensis Jacq.) Using Temporary Immersion System

Tissue culture technique has been used to produce elite oil palm clonal materials in commercial scale. Proliferation of embryogenic callus in oil palm commonly utilizes solid and liquid medium, but the result are still not optimum. Recently, researchers have developed an improved technique called Temporary Immersion System (TIS) and this technique has shown great potential to increase proliferation rate of the embryogenic callus. This research can be divided into two groups i.e. selection of liquid medium using shaker to increase the proliferation rate and proliferation of embryogenic callus using TIS. The objectives of this research are to obtain the best medium for increasing embryogenic proliferation rate using shaker and to obtain the best medium and time interval of immersion to increase the proliferation rate using TIS. Approximately 0.3g of embryogenic callus from embryoid-line A1 were weighted and cultured in twenty two types of liquid medium for the first research and two types of medium with the composition of MSD and MSA for the second research. The proliferation rate based on fresh weight obtained from week 8 observations. The data was analysed with Anova of F (α = 5%) and then with DMRT in α = 5%. TIS observation was done every 1h, 3h, 6h of immersion time with time interval of 3min. The results showed that the best medium composition to increase proliferation of embryogenic callus using shaker was MSA9, the best medium composition to increase proliferation of embryogenic callus using TIS was MSD, and the best time interval of immersion was every 3h for 3min.

Evaluation of a new vessel system based on temporary immersion system for micropropagation

The aim of this study was to evaluate a new culture vessel Plantform bioreactor, based on the temporary immersion system principle for micropropagation of Digitalis lutea×purpurea, Echinacea purpurea and Rubus idaeus. The multiplication ratio and shoot quality in bioreactor were either similar to or better than these on agar medium. The shoot number was similar in both systems for Digitalis and Rubus, while Echinacea had a significantly higher shoot number of good quality in bioreactor. Digitalis and Echinacea gained more fresh weights in bioreactor, while more for Rubus on agar medium. However, the dry weight was similar between the two systems for all three species. Changes in carbohydrate were evident for all species. Autoclaving media could breakdown sucrose, which is both pH- and nutrient-dependent. This study has demonstrated that the Plantform bioreactor is suitable for plant micropropagation.

The Evaluation of Children with an Autism Spectrum Disorder: Adaptations to Accommodate a Telepractice Model of Clinical Care

Telepractice is increasingly applied to assessment and intervention for persons with autism spectrum disorders (ASD), and the use of telepractice for delivering augmentative and alternative communication (AAC) services is expanding. To date, no studies have focused on the assessment of communication in children with ASD who are minimally verbal. This case exploration represents a first attempt to apply the clinical assessment approach (also known as Visual Immersion System) developed in the Autism Language Program in the Center for Communication Enhancement (CCE) at Boston Children's Hospital (BCH) through a telepractice service delivery model. In this paper we detail an on-site evaluation, and then describe the application of the various evaluation components to telepractice delivery. Our findings provide preliminary evidence that communication assessment information can be successfully obtained for this population via telepractice. The results demonstrated that the telepractice evaluation took greater time to complete than the on-site evaluation, likely due to both technical and cultural factors. Further study is warranted particularly in the delivery of family instruction materials prior to the evaluation and to quality assurance measures.

Leucojum aestivum L. in vitro bulbs induction and acclimatization

Abstract The effects of growth retardants (paclobutrazol and ancymidol), sucrose, GA3 (gibberellic acid) and physical state of the medium (solid and liquid — Rita® temporary immersion system) on in vitro induction of Leucojum aestivum bulbs and their acclimatization were studied. Paclobutrazol, regardless of the physical state of the medium, stimulated the formation of bulbs (99.3%). Under the influence 90 g L−1 of sucrose or paclobutrazol the bulbs with the highest fresh weight (FW) were formed (250 mg and 208.8 mg, respectively). However, the addition of ancymidol to the liquid medium led to obtaining the bulbs showing the highest number of leaves and roots (63.2% and 91.7%, respectively). The scanning microscopy study proved that plants obtained in the medium containing GA3 produced the stomata which most closely resembled to the one observed in the mother plant. Cytometric analysis of all regenerants revealed absence of changes in the nuclear DNA content. The maximum survival rate (100%) was observed for plants derived from liquid medium containing 90 g L−1 of sucrose. Somewhat fewer plants were acclimatized after their cultivations in liquid medium enriched with paclobutrazol or ancymidol. The temporary immersion system led to perform successful ex vitro adaptation of Leucojum aestivum plants.

Xanthone, benzophenone and bioflavonoid accumulation in Cyclopia genistoides (L.) Vent. (honeybush) shoot cultures grown on membrane rafts and in a temporary immersion system

In vitro shoots of the commercially important South-African legume Cyclopia genistoides were grown in different liquid culture systems [membrane rafts (MR) and a temporary immersion bioreactor (TIB)] and evaluated for the accumulation of phenolic secondary metabolites. The major constituents of the investigated cultures were medicinally relevant xanthones [mangiferin (M) and isomangiferin (IM)] and benzophenone derivatives [iriflophenone 3-C-β-glucoside (IG)]. The highest concentrations of M, IM and IG in MR-grown shoots were 1,843.59, 712.02 and 594.29 mg 100 g−1 dry wt, respectively. Bioreactor cultivation provided higher peak concentrations of M (2,622.70 mg 100 g−1 dry wt), IM (757.40 mg 100 g−1 dry wt) and IG (648.30 mg 100 g−1 dry wt) which corresponded to the respective productivities of 5.48, 1.58 and 3.04 mg l−1 d−1. The results indicate that TIB cultures of C. genistoides may be utilized as an alternative source of the above constituents, particularly IM and IG, which are relatively expensive and so far hardly available from commercial sources.

The Relationship between English and Persian Phonological Awareness, Rapid Autamatized Naming and Students’ Reading Achievement in Partial Immersion and Non-Immersion Programs

The cognitive predictors (i.e.,Phonological Awareness, and Rapid Automatized Naming) underlying reading achievement have not been researched in Iranian partial English immersion and non-immersion programs. The present study sought to investigate the relationship between English and Persian Phonological Awareness (PA), Rapid Autamatized Naming (RAN) and reading achievementof Iranian students in partial immersion and non-immersion programs. To this end, one hundred forty five students from three different grade levels in a partial English immersion program and 95 students from three different grade levels in a non-immersion program were chosen. Six different English and Persian tests were utilized (namely, the Cambridge English for Young Learners (YLE) test for Reading,the Persian reading achievement test, the English and Persian Phonological AwarenessSound Detection tests, and the English and Persian Rapid Automatized Naming Tests). Given the design of the study, a number of statistical tests were run. The main findingswere as follows: learners’ reading achievement could significantly be predicted through both English and Persian PA and RAN.Furthermore, learning English in a partial English immersion system improves learners’ reading achievement and cognitive predictors compared with non-immersion program. The findings suggest that by teaching learners PA and RAN skills, their reading achievement improves in both English and Persian.

Improved propagation of vanilla (Vanilla planifolia Jacks. ex Andrews) using a temporary immersion system

Here, we evaluated the efficiency of shoot multiplication of Vanilla planifolia Jacks. ex Andrews using solid medium, partial immersion, and a temporary immersion system (TIS) to improve micropropagation in this species. Clusters of shoots were cultivated in vitro using Murashige and Skoog (MS) medium supplemented with 9.55 μM benzyladenine (BA) and 100 mL L−1 coconut water. For the TIS, a RITA® system was used and three immersion frequencies were evaluated (every 4, 8, and 12 h) with an immersion time of 2 min. After 30-d culture, the TIS produced the maximum multiplication rate (14.27 shoots per explant) when using an immersion frequency of 2 min every 4 h, followed by the partial immersion system (8.64 shoots per explant), and solid medium (5.80 shoots per explant). Next, the effect of the volume of culture medium per explant was also evaluated for TIS. The most suitable volume of culture medium for shoot formation was 25 mL per explant, which increased the rate of multiplication to 17.54 shoots per explant. Root initiation was 90% successful in TIS using half-strength MS medium supplemented with 0.44 μM naphthaleneacetic acid (NAA) and an immersion frequency of 2 min every 4 h. With this system, the shoot multiplication rate increased threefold compared to that obtained with solid medium. In addition, this system produced good results for the transplantation and acclimation (90% of survival) of in vitro-derived plants. These results offer new options for large-scale micropropagation of vanilla.

Large-Scale Continuous Immersion System for Layer-by-Layer Deposition of Flame Retardant and Conductive Nanocoatings on Fabric

A continuous immersion system has been developed to deposit multifunctional, multilayer thin films using layer-by-layer assembly. An initial demonstration of this device is presented here by coating cotton fabric, a complex three-dimensional substrate, with a flame-retardant recipe composed of chitosan (CH) and poly(sodium phosphate) (PSP) and a conductive recipe composed of poly(diallyldimethylammonium chloride) and multiwalled carbon nanotubes stabilized with sodium deoxycholate. Electron microscopy images reveal an unaltered weave structure for the conductive coating and the formation of a CH/PSP protective thermal barrier on fabric coated by the machine. Flame retardant and conductivity measurements of continuously coated fabric achieve properties comparable to handmade samples, and the large-scale coater showed a high level of consistency. This unique device provides an important proof-of-concept and lays the groundwork for scale-up to an industrial-scale system that could be used to produce these na...

Micropropagation of the pistachio and its rootstocks by temporary immersion system

Although several studies have been reported on the micropropagation of the pistachio and its rootstocks, to date none of them had been efficient on the mass production of these plants in bioreactor systems. Thus, the micropropagation of juvenile pistachio shoot tips and nodal buds was investigated in a temporary immersion bioreactor system (RITA®) and on a conventional semi-solid medium. Among the tested immersion conditions, immersion for 24 min every 16 h reduced vitrification and improved proliferation in the pistachio. Interactions were evident in immersion time and frequency in nodal segments. Nodal buds were better than shoot tips as the highest multiple shoot formation was recorded in MS medium containing 4 mg L−1 BA and 0.1 mg L−1 GA3 in RITA®. Although shoot tip necrosis (STN) was observed in shoots proliferated on semi-solid MS medium, such a symptom did not occur in shoots sprouted in the RITA®. Additionally, these optimized conditions were applied to nodal buds of mature male pistachio ‘Atli’ and Pistacia rootstocks (P. khinjuk Stocks and P. atlantica Desf.), and the micropropagation in the bioreactor system, in comparison to the semi-solid medium, was also improved. Furthermore, in vitro rooting of pistachio plantlets, despite the lower range (27.5 %), was also achieved in RITA®. However, rooting was better on semi-solid medium for all tested species (ranged between 50 and 70 %). The results of this study showed that RITA® could be used for the mass propagation of pistachio and its rootstocks, as well as for other woody plant species.

Effect of BA Treatments on Morphology and Physiology of Proliferated Shoots of <i>Bambusa vulgaris</i> Schrad. Ex Wendl in Temporary Immersion

Axillary buds, collected from greenhouse-grown plants of Bambusa vulgaris Schrad. ex Wendl (B. vulgaris), were incubated on a static liquid culture medium, Murashige and Skoog (MS) medium with 2% (w/v) sucrose, and supplemented with 12.0 μM 6-benzyladenine (BA). They were transferred to a temporary immersion system (TIS) using liquid MS medium supplemented with 0 (CK-free medium), 6.0, 12.0, 18.0 μM BA. The morphological and anatomical indicators were measured. The BA influenced in vitro multiplication of B. vulgaris. The best results were achieved in the SIT with a concentration of 6.0 μM of BA, which increased the number of shoots (5.1 shoots/explant) in the absence of hyperhydric shoots. Results demonstrated that the water content in the sprouts increased with 12.0 and 18.0 μM BA every four hours. Furthermore, these high levels of BA contributed to a lower accumulation of phenolic compounds and lignin content. The total chlorophyll significantly increased when using 6.0 uM BA, but decreased both parameters with other treatments. These results favor to increase the number of shoots/explants during in vitro multiplication. They will also optimize the in vitro culture conditions, leading to an improvement of in vitro propagation methods for this species.

Genetic transformation of hop (Humulus lupulus L. cv. Tettnanger) by particle bombardment and plant regeneration using a temporary immersion system

The efficiency of micropropagation of double-node shoots of hop (Humulus lupulus L. cv. Tettnanger) was evaluated using semi-solid and liquid culture medium in RITA® temporary immersion bioreactors. The highest fresh and dry weight of shoots, average number of shoots, and multiplication rate were obtained using the RITA® system, whereas the longest shoots were obtained on semi-solid medium. Moreover, shoot length was affected significantly by the inoculum density of double-node shoots in RITA® vessels. In addition, the RITA® bioreactors were suitable for shoot induction from organogenic calli. The percentage of shoot induction and the shoot fresh and dry weights were significantly higher in the RITA® system than in semi-solid medium. The age of organogenic calli and inoculum density significantly affected the induction of shoots from organogenic calli. The optimum conditions for DNA delivery into hop organogenic calli using the biolistic particle delivery system were also determined. Organogenic calli were bombarded with the plasmid pSR5-2 (gusA and nptII) varying helium pressure (900, 1,100, or 1,350 psi) and target distance (6, 9, or 12 cm). The highest gusA transient activity was obtained using a pressure of 900 psi and a target distance of 6 cm. For stable genetic transformation, 3-wk-old organogenic calli were bombarded with the plasmid pCAMBIA1303 (gusA, mgfp5, and hpt) using these optimum conditions. Stable gusA expression was observed in organogenic calli and shoots after 4 wk of culture on selection medium containing 2.5 mg l−1 hygromycin. The presence of the mgfp5 gene in the hop genome was confirmed by PCR.

Overexpression of the chestnut CsTL1 gene coding for a thaumatin-like protein in somatic embryos of Quercus robur

Somatic embryos of Quercus robur derived from mature and juvenile trees were transformed with the CsTL1 gene coding for an antifungal protein. Clumps of somatic embryos were inoculated with Agrobacterium tumefaciens strain EHA105 harbouring a binary vector (pKWG2D-TAU) that included the green fluorescence protein (GFP) reporter gene. After 24 weeks of culture on selective medium, significantly higher transformation frequencies were obtained in a temporary immersion system (TIS) than in semi-solid medium (ranging from 1.4 to 9.6 % for the four genotypes). The first transformants appeared after 10 weeks and GFP activity was evaluated in all kan-resistant embryogenic lines. The transgenic nature of lines was confirmed by PCR and Southern blotting. CsTL1 transcripts were detected in all transgenic lines (TL) analyzed by quantitative real-time PCR, and levels of expression were significantly higher than in the wild type (wt), reaching up to 24 times higher in some genotypes. Sixty-two stable TL were obtained (38 derived from centenarian trees). Plant conversion rates of transformed somatic embryos were similar to those of the wt and were increased by use of the TIS. GFP fluorescence was detected in roots and shoots of transgenic plants, which confirmed stable transformation of regenerants.

Global DNA methylation profiles of somatic embryos of peach palm (Bactris gasipaes Kunth) are influenced by cryoprotectants and droplet-vitrification cryopreservation

Regrowth capacity and genetic stability of plants recovered following cryopreservation are associated with changes in DNA epigenetics, particularly in DNA methylation levels. In this study, global DNA methylation profiles associated with frequency of regrowth of peach palm (Bactris gasipaes) somatic embryos following cryopreservation using droplet-vitrification were investigated. Somatic embryo clusters (SEC) subjected to plant vitrification solution 3 (PVS3) for different durations (0, 60, 120, 180, and 240 min) were evaluated for regrowth capacity. The highest frequency of regrowth (52.4 %) was obtained when SEC were incubated in PVS3 for 120 min prior to droplet-vitrification cryopreservation. Global DNA methylation profiles were influenced by both cryoprotectants and droplet-vitrification cryopreservation. Incubation of SEC in PVS3 for limited durations not only reduced frequency of regrowth, but also increased DNA methylations levels when compared with proliferating SEC grown in a temporary immersion system. Although SEC subjected to cryopreservation exhibited the highest DNA methylation variation, 120 min SEC incubation in a PVS3 solution resulted in the recovery of initial global methylation profiles after 24 weeks of regrowth.

High-efficiency Agrobacterium-mediated transformation in Quercus robur: selection by use of a temporary immersion system and assessment by quantitative PCR

An efficient protocol for genetic transformation of somatic embryos of Quercus robur by selection in a temporary immersion system is reported. The transformation frequency was 5 times higher than achieved by conventional culture on semi-solid medium, ranging between 6 and 26 % for the four genotypes evaluated. Clumps of globular or torpedo somatic embryos were precultured for 7–10 days, inoculated with Agrobacterium tumefaciens strain EHA105:p35SGUSINT and cocultivated for 4 days before being cultured for 4 weeks on semi-solid selection medium supplemented with 25 mg L−1 kanamycin. Explants were transferred to RITA® bioreactors and subjected to a two-step selection protocol involving immersion in liquid medium supplemented with 25 mg L−1 kanamycin, for 18 weeks, and then with 75 mg L−1 kanamycin. Putatively transformed explants appeared after serial transfer to selection medium over 12–16 weeks. The presence of neomycin phosphotransferase II and β-glucuronidase genes in the plant genome was confirmed by histochemical and molecular analysis, and the copy number was determined by Southern blotting and real-time quantitative polymerase chain reaction. Transformed somatic embryos were germinated and transferred to soil for acclimatization, approximately 8 months after inoculation of the original tissue with bacteria. As the limiting factor for recovery of plants from oak embryogenic lines is the low embryo conversion rate, axillary shoot lines were established from transformed germinated embryos. Transformed embryos and shoots were cultured in medium with or without kanamycin and the responses to several morphogenetic processes (recovery after cryopreservation, germination, shoot proliferation, and rooting) were evaluated.

Elicitation of galanthamine production by leucojum aestivum shoots grown in temporary immersion system

The influence of different elicitors (copper sulfate, silver nitrate, salicylic acid and methyl jasmonate), on both the growth and alkaloid production of Leucojum aestivum shoots grown in a temporary immersion system was studied. Seven Amaryllidaceae alkaloids and three protoalkaloids were quantitatively determined by GC-MS analysis in leaves and bulblets, separately. Methyl jasmonate was found to significantly improve the production of galanthamine (GAL) in both leaves and bulblets. The content of GAL released to the liquid nutrient medium was also measured. The release of GAL into the liquid medium took place mainly in the first 2 weeks determined by harvesting the liquid nutrient medium after 2 weeks and measuring the GAL content (1st subculturing step).

Nodule cluster cultures and temporary immersion bioreactors as a high performance micropropagation strategy in pineapple (Ananas comosus var. comosus)

Biotechnology tools based on tissue culture techniques allow mass clonal propagation of plants with desired genetic and sanitary features. In the present work a pineapple micropropagation protocol was established based on nodule cluster cultures (NC) associated with different culture systems. Leaf segments from in vitro-grown plantlets cultivated in culture medium supplemented with 2μM NAA and 8μM 2-iP or BAP resulted in the highest rates of NC induction. NC fresh weight increased ratio was more efficient in a culture medium supplemented with 2μM NAA and 2μM BAP in twin-flasks temporary immersion system (TIS-TF). This culture medium was also efficient for the microshoots development in RITA® temporary immersion system (TIS-RITA®). Microshoots elongation in permanent immersion system (PIS) was more efficient in a culture medium supplemented with 10μM GA3. Shoots longer than 2cm showed 92% survival rate when acclimatized. Free polyamine endogenous levels was measured and showed higher contents for the TIS-TF, followed by TIS-RITA®, and PIS, respectively. Putrescine levels also followed this pattern, being absent in PIS, which, on the other hand, showed higher levels of spermine and spermidine. The results indicate the establishment of efficient protocol for pineapple micropropagation, with a high multiplication and regenerative rates and great potential for agriculture application. The biochemical analysis suggests that besides culture medium composition, flask design and culture system also drastically affects the plant's metabolism toward multiplication and plant regeneration.

A comparative study between solid and liquid cultures relative to callus growth<br>and somatic embryo formation in date palm (Phoenix dactylifera L.) cv. Zaghlool

Tissue culture techniques enable mass propagation of elite cultivars of date palm (Phoenix dactylifera L.). The main limitations for date palm in vitro multiplication are the low rates achieved using solidified medium and the long period needed to produce acclimatized plantlets. This research focuses on the comparison between different culture types and plant growth regulator (PGR) combinations on callus growth and somatic embryo formation of cv. Zaghlool. For callus growth, 200 mg friable embryogenic callus dispensed in Rasotherm and Phytacon flasks containing 200 ml liquid (100 rotations per minute) and in temporary immersion system, RITA ® bioreactor (5 min immersion every 12 h), were compared with cultures grown on 200 ml solidified medium. For somatic embryo formation, 500 mg of friable embryogenic callus grown in Erlenmeyer flasks filled with 50 ml liquid or solid MS medium. The medium was supplemented with 0.1 mg l -1 Naphthaleneacetic acid (NAA), 1.5 g l -1 activated charcoal (AC) with or without 0.05 mg l -1 6-Benzyl amino purine (BAP) compared to PGR-free medium. Results proved that cell suspension cultures produced the highest callus fresh mass as compared to the other systems tested, and the callus fresh mass reached 4 g after 16 weeks. The temporary immersion system did not significantly enhance the fresh mass of callus compared to the solidified medium. For somatic embryo induction, the number of somatic embryos increased in cell suspensions 6-16 fold compared to the solidified medium. Using liquid MS medium enriched with 0.1 mg l -1 NAA and 1.5 g l -1 AC gave rise to the highest number of somatic embryos formed from 500 mg initial callus: 160 embryos. The number of somatic embryos was also affected by the callus source. The calli induced from leaflet segments excised from converted somatic embryos resulted in a lower somatic embryo number than those of shoot tip origin with about 60 somatic embryos per 500 mg callus. The formation of somatic embryos using liquid medium required only 6 weeks, thus considerably reducing the previously reported period of 18 weeks which is required for somatic embryo formation using solidified media.

ESTABLISHMENT OF A SHOOT PROLIFERATION PROTOCOL FOR BANANA (ABB GROUP) CV. ‘PISANG AWAK’ VIA TEMPORARY IMMERSION SYSTEM

□ Initial study of evaluating the factors affecting shoot proliferation of ‘Pisang Awak’ using temporary immersion system was carried out using four half shoots in each experimental unit as explants and repeated with four to seven experimental units for each factor. The inoculum size finally used for shoot proliferation was eight half-shoots for each experimental unit (250 mL culture flask) in MS liquid medium supplemented with 5.0 mg L−1 benzyladenine (BA) and 30 g L−1 sucrose, the shoot proliferation medium and an average of 4.9 ± 0.3 shoots was produced from each half-shoot explant after five weeks of culture. The final protocol for shoot proliferation using the temporary immersion system was established by inoculating eight half-shoot explants into the shoot proliferation medium adjusted to pH 5.7 with the inoculated shoots immersed once a day in the medium for 10 minutes. The cultures were incubated under continuous light for enhancement of shoot proliferation without vitrification.

The status of temporary immersion system (TIS) technology for plant micropropagation

The development of temporary immersion systems (TIS) for in vitro culture, in the late 1990s heralded new and semi-automated means of micropropagation of plants for agricultural and conservation purposes. By 2005, various systems had been developed and tested and their benefits were proven with a variety of commercially-important crops such as sugar-cane, coffee, banana, and yam. Progress in the field and the adoption of these systems in subsequent years has been reviewed. Improvements on previously-developed TIS protocols for economically-important species have been reported, and this continues to attest to their benefits when compared with protocols that employ semi-solid or liquid media. Though TIS protocols have recently been developed for other species, this list is limited. On the other hand, there appears to be an increasingly great interest in TIS technology for the production of secondary metabolites and for physiological studies.