The Development of In Vitro Culture Sterilization Method of Gametophyte Explant Lopholejeunea sp.

In vitro cultures of leafy liverworts are still facing significant challenges due to high-level of explant contamination. The sterilization process can easily damage the structure of liverwort after exposure to the disinfectant. This study was to determine the concentration and time exposure of commercial bleach as a disinfectant to suppress contamination using the gametophyte culture of Lopholejeunea sp. The experiment consisted of control and six treatment combinations of commercial bleach with concentration 0.5, 0.75, and 1% (v/v), and exposure time (60 and 90 seconds). The type and location of contamination, the color of the explants after sterilization, and response after 30 days were observed. The results showed that the 0.75% bleach with 60 and 90 seconds exposure time had a lower contamination until the 7th day of culture. The most common type of contamination is bacteria and fungi that arise from the explant. Despite the contamination, it did not inhibit shoot formation. Further studies still needed to determine the type of fungicides and antibiotics with the most potent concentration and exposure time should be tested to obtain an axenic and viable culture of liverworts Lopholejeunea sp.

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The Development of Gametophyte Sterilization Method for Liverworts Acrolejeunea fertilis (Reinw., Blume and Nees) Schiffn. In vitro Culture

HAYATI Journal of Biosciences ◽

10.4308/hjb.28.1.39 ◽

2021 ◽

Vol 28 (1) ◽

pp. 39

Author(s):

Mouleidi Dwi Putri ◽

Windri Handayani ◽

Astari Dwiranti ◽

Andi Salamah ◽

Niarsi Merry Hemelda ◽

...

Keyword(s):

In Vitro Culture ◽

Exposure Time ◽

Common Type ◽

Internal Contamination ◽

Sterilization Method ◽

Combination Treatments ◽

Sterilization Process ◽

Bacteria And Fungi ◽

High Level

In vitro culture gametophytes of leafy liverworts often have problems in their sterilization process. These problems due to the high level of contamination and the fragile structure of the gametophyte leafy liverworts. The structures can be easily to damage after exposure to disinfectant. This study aimed to observe the concentration and the exposure time of “Bayclin” commercial bleach to suppress contamination with the viability of Acrolejeunea fertilisgametophytic explants. This research was conducted using control and 6 combination treatments with “Bayclin” concentration (1.00%, 1.25%, and 1.50%) and exposure time 60 and 120 seconds, then accompanied by the addition of Tetracycline 2.5 mg/ml. The qualitative parameters observed were the explant color, the type and location of contamination, and the growth of explants. The quantitative parameters were the percentage of contamination, the percentage of growth, and the number of new branches. The results showed that “Bayclin” 1.25% and 1.50% with 60 seconds exposure time has the lowest percentage of contamination which is 70% until the 7th days after planting. The most common type of internal contamination from the explant is bacteria and fungi. However, the growth of the new branch still occurs in some explants even though it has been contaminated and browned.

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EFEKTIVITAS MERKURI KLORIDA (HgCl2) PADA STERILISASI TUNAS SAMPING JATI (Tectona grandis) IN VITRO

Jurnal Bioteknologi & Biosains Indonesia (JBBI) ◽

10.29122/jbbi.v4i2.2540 ◽

2017 ◽

Vol 4 (2) ◽

pp. 25

Author(s):

Yusuf Sigit Ahmad Fauzan ◽

. Supriyanto ◽

Teuku Tajuddin

Keyword(s):

In Vitro Culture ◽

Tectona Grandis ◽

In Vitro Cultures ◽

Mercury Chloride ◽

Main Obstacle ◽

Sterilization Process ◽

High Level ◽

Growth Of Fungi ◽

Tectona Grandis L.F

Effectiveness of Mercury Chloride (HgCl2) in Sterilization of Teak (Tectona grandis L.f.) In VitroThe main obstacle in obtaining sterile materials in in vitro cultures derived from meristems is high level of surface contamination caused by fungi and bacteria, which often results in explant death. The objective of this study was to obtain an appropriate mercury chloride (HgCl2) concentration for the sterilization of Tectona grandis nodes in in vitro culture. One cm long-sized nodes with 0.2 mm diameter were immersed in HgCl2at concentrations of 0, 100, 200 and 300 mg/L for 3 minutes. The results showed that the higher concentration of HgCl2was able to suppress the growth of fungi and bacteria and increased the percentage of aseptic explants. The best HgCl2concentration was 300 mg/L since it suppressed the growth of fungi and bacteria up to 100% and 75%, respectively, and produced the highest aseptic explants of 85% at 9 days after treatment. The small sized explants supported the sterilization process and reduced browning levels.Keywords: Browning, HgCl2, in vitro, sterilization, T. grandisABSTRAKKendala utama dalam mendapatkan material steril pada kultur in vitro yang berasal dari meristem adalah tingginya tingkat kontaminasi permukaan yang disebabkan oleh jamur dan bakteri, dan sering menyebabkan kematian eksplan. Tujuan penelitian ini adalah untuk memperoleh konsentrasi merkuri klorida (HgCl2) yang tepat untuk sterilisasi eksplan tunas samping tanaman jati (Tectona grandis) pada kultur in vitro. Tunas samping berukuran 1 cm dan diameter 0,2 mm direndam dalam HgCl2 pada konsentrasi 0, 100, 200 dan 300 mg/L selama 3 menit. Hasil penelitian menunjukkan bahwa penambahan konsentrasi HgCl2 yang semakin tinggi mampu menekan pertumbuhan jamur dan bakteri pada eksplan serta meningkatkan persentase eksplan aseptik. HgCl2 dengan konsentrasi 300 mg/L merupakan konsentrasi terbaik karena dapat menekan pertumbuhan jamur hingga 100% dan bakteri mencapai 75%, serta menghasilkan tingkat eksplan aseptik dan hidup tertinggi yaitu sebesar 85% pada 9 hari setelah perlakuan. Ukuran eksplan yang kecil mendukung proses sterilisasi dan mengurangi tingkat browning. Kata kunci: HgCl2,in vitro, pencoklatan jaringan, sterilisasi, T. grandis, Received: 02 November 2017 Accepted: 14 December 2017 Published: 29 December 2017

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Introduction of hyssopus officinalis l . into in vitro culture to optimize the conditions for obtaining callus tissues and microclonal propagation as a promising metod of innovative agrobiotechnologies

BIO Web of Conferences ◽

10.1051/bioconf/20213005006 ◽

2021 ◽

Vol 30 ◽

pp. 05006

Author(s):

Elena Maslova ◽

Natalya Gulya ◽

Tatyana Perelugina ◽

Valeria Semykina ◽

Elena Kalashnikova

Keyword(s):

In Vitro Culture ◽

Exposure Time ◽

Crop Production ◽

Callus Tissue ◽

Nutrient Media ◽

Sterilization Process ◽

Callus Tissues ◽

Active Substances ◽

Hyssopus Officinalis

The sterilization process of plant explants of H. officinalis was optimized when introduced into an in vitro culture, the most effective sterilization modes, optimal sterilizing agents, their exposure time and concentration were selected. Callus tissues and mini-plants of H. officinalis were obtained in vitro and the most optimal nutrient media were determined both for microclonal propagation and for the induction of callus tissue H. officinalis, which can be further used for mass cultivation of cell and culture and obtaining safe bio-additives with active substances for livestock and crop production as a part of the development of modern agrobiotechnologies.

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Photodynamic Therapy Combined with Antibiotics or Antifungals against Microorganisms That Cause Skin and Soft Tissue Infections: A Planktonic and Biofilm Approach to Overcome Resistances

Pharmaceuticals ◽

10.3390/ph14070603 ◽

2021 ◽

Vol 14 (7) ◽

pp. 603

Author(s):

Vanesa Pérez-Laguna ◽

Isabel García-Luque ◽

Sofía Ballesta ◽

Antonio Rezusta ◽

Yolanda Gilaberte

Keyword(s):

Photodynamic Therapy ◽

Soft Tissues ◽

Combined Treatment ◽

Resistance Pattern ◽

Antimicrobial Photodynamic Therapy ◽

Antimicrobial Drugs ◽

Bacteria And Fungi ◽

High Level ◽

Selection Of

The present review covers combination approaches of antimicrobial photodynamic therapy (aPDT) plus antibiotics or antifungals to attack bacteria and fungi in vitro (both planktonic and biofilm forms) focused on those microorganisms that cause infections in skin and soft tissues. The combination can prevent failure in the fight against these microorganisms: antimicrobial drugs can increase the susceptibility of microorganisms to aPDT and prevent the possibility of regrowth of those that were not inactivated during the irradiation; meanwhile, aPDT is effective regardless of the resistance pattern of the strain and their use does not contribute to the selection of antimicrobial resistance. Additive or synergistic antimicrobial effects in vitro are evaluated and the best combinations are presented. The use of combined treatment of aPDT with antimicrobials could help overcome the difficulty of fighting high level of resistance microorganisms and, as it is a multi-target approach, it could make the selection of resistant microorganisms more difficult.

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MTMS-Based Aerogel Constructs for Immobilization of Plant Hairy Roots: Effects on Proliferation of Rindera graeca Biomass and Extracellular Secretion of Naphthoquinones

Journal of Functional Biomaterials ◽

10.3390/jfb12010019 ◽

2021 ◽

Vol 12 (1) ◽

pp. 19

Author(s):

Bartosz Nowak ◽

Mateusz Kawka ◽

Kamil Wierzchowski ◽

Katarzyna Sykłowska-Baranek ◽

Maciej Pilarek

Keyword(s):

Polyurethane Foam ◽

Bioactive Compounds ◽

Hairy Roots ◽

Reference System ◽

Culture System ◽

In Vitro Cultures ◽

Fresh Mass ◽

Extracellular Secretion ◽

High Level

Unique biosynthetic abilities revealed by plants determine in vitro cultures of hairy roots as a suitable source of pharmaceutically relevant bioactive compounds. The basic aim of the study was to examine the applicability of aerogel composed of methyltrimethoxysilane (MTMS) for immobilization of Rindera graeca hairy roots by identifying quantitative effects of biomass proliferation and naphthoquinones extracellular secretion in the aerogel-supported culture system. R. graeca hairy roots were simultaneously cultured for 28-days, as (i) nonimmobilized biomass (reference system), (ii) biomass immobilized on macroporous polyurethane foam (PUF), (iii) biomass with disintegrated MTMS aerogel, (iv) biomass immobilized on polypropylene (PP) fibers (as control), and (v) biomass immobilized on monolithic PP-reinforced MTMS aerogel. MTMS aerogel exhibited high level of biocompatibility toward R. graeca hairy roots which grew into the structure of monolithic aerogel-based constructs. Monolithic MTMS-based constructs significantly promoted the proliferation of hairy roots, resulting in 55% higher fresh mass than the reference system. The highest level of naphthoquinones productivity, i.e., 653 µg gDW−1, was noted for PUF-supported culture system.

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Optimasi sterilisasi permukaan eksplan stek mikro tanaman karet Optimization of surface sterilization on rubber microcutting explan

E-Journal Menara Perkebunan ◽

10.22302/iribb.jur.mp.v81i1.55 ◽

2016 ◽

Vol 81 (1) ◽

Author(s):

Irfan MARTIANSYAH ◽

Deden Dewantara ERIS ◽

. NURHAIMI-HARIS ◽

Darmono TANIWIRYONO

Keyword(s):

Primary Culture ◽

Second Step ◽

Contamination Level ◽

Surface Sterilization ◽

Major Constraint ◽

Culture Stage ◽

Sterilization Process ◽

High Level

AbstractAn increasing number of explants is necessary toobtain plantlets in large quantities, for mass propagationof rubber plants. However, high level of contamination atthe primary culture stage is still a major constraint in invitro microcutting of rubber. The aim of this study was tooptimize surface sterilization procedures to reduce micro-bial contamination at the primary culture. Sterilizationexperiment was conducted in two step., The first step wasto determine the effect of washing the explants withrunning water prior to sterilization and then using Deso-germe, ethanol or H 2 O 2 , while the second step was toidentify the suitable sterilization process on reducing thelevel of contamination. The results showed that the surfacesterilization with only one type of sterilization agent couldnot reduce contamination level caused either by bacteriaor fungi, while sterilization with three types of sterilizingagents increased the number of dead explants. The besttreatment for surface sterilization was the directsterilization of explants using 70% ethanol for one minuteand 17.6% H 2 O 2 for 20 minutes without washing with tapwater (A-CD treatment). The percentage of viable andaseptic explantsof this treatment was 76.7%, which wassignificantly higher than those of other treatments. Thistreatment reduced contamination level to 21.7%.AbstrakPeningkatan jumlah eksplan sangat diperlukan untukmemperoleh planlet dalam jumlah besar pada perbanyakanmassal tanaman karet secara in vitro. Namun, tingginyatingkat kontaminasi pada tahap kultur primer masih me-rupakan kendala utama dalam kultur stek mikro tanamankaret. Tujuan penelitian adalah mengoptimasi prosedursterilisasi permukaan eksplan untuk mengurangi jumlaheksplan yang terkontaminasi mikroba pada tahap kulturprimer. Percobaan sterilisasi dilaksanakan dalam duatahap, tahap pertama untuk mengetahui pengaruh pen-cucian eksplan dengan air mengalir pada awal sterilisasiserta penggunaan Desogerme, etanol dan H 2 O 2 , sedang-kan tahap kedua untuk mendapatkan proses sterilisasi yangpaling sesuai dalam menurunkan tingkat kontaminasi.Hasil penelitian menunjukkan bahwa perlakuan sterilisasipermukaan yang menggunakan satu jenis bahan sterilantidak dapat mengurangi kontaminasi, baik oleh bakterimaupun cendawan. Perlakuan sterilisasi eksplan dengantiga jenis bahan sterilan meningkatkan kematian eksplan.Perlakuan sterilisasi permukaan terbaik adalah sterilisasilangsung eksplan menggunakan etanol 70% selama satu menit dan H 2 O 2 17,6% selama 20 menit, tanpa pencuciandengan air mengalir (perlakuan A-CD). Persentase eksplansteril yang hidup sebesar 76,7%, berbeda nyata dibanding-kan dengan perlakuan lainnya. Perlakuan tersebut dapatmengurangi kontaminasi menjadi sebesar 21,7%.

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883 PB 279 REGENERATING ADVENTITIOUS PLANTS FROM IN VITRO CULTURE OF LIATRIS SPICATA (L.) WILLD. COTYLEDONS

HortScience ◽

10.21273/hortsci.29.5.560d ◽

1994 ◽

Vol 29 (5) ◽

pp. 560d-560

Author(s):

Dennis P. Stimart ◽

John C. Mather

Keyword(s):

In Vitro Culture ◽

Adventitious Shoots ◽

Adventitious Shoot ◽

Shoot Formation ◽

Ms Medium ◽

Ex Vitro ◽

Adventitious Shoot Formation ◽

Murashige Skoog ◽

Micropropagated Plants

Cotyledons from developing embryos 6 to 8 weeks old of Liatris spicata (blazing star) were cultured on Murashige-Skoog (MS) medium containing 0, 0.4, 4.4, and 44.4 μ M benzyladenine (BA) or 0, 0.2, 2.2, and 22.2 μ M thidiazuron (TDZ) to induce adventitious shoot formation. The highest percent of cotyledons forming shoots with highest shoot counts was on medium containing 2.2 μ M TDZ. Vitreous shoots formed on medium with 22.2 μ M TDZ. Callus derived from cotyledons and cultured on medium containing 4.44 μ M BA or 2.2 μ M TDZ formed adventitious shoots with highest shoot counts on 4.44 μ M BA. Adventitious shoots derived from cotyledons and callus were rooted on MS medium with 5.0 μ Mindole-3-butyric acid, acclimatized and grown ex vitro. All micropropagated plants appeared similar to each other.

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Regenerating Adventitious Shoots from in Vitro Culture of Liatris spicata (L.) Willd. Cotyledons

HortScience ◽

10.21273/hortsci.31.1.154 ◽

1996 ◽

Vol 31 (1) ◽

pp. 154-155

Author(s):

Dennis P. Stimart ◽

John C. Mather

Keyword(s):

In Vitro Culture ◽

Adventitious Shoots ◽

Adventitious Shoot ◽

Shoot Formation ◽

Ms Medium ◽

Adventitious Shoot Formation ◽

Regenerated Plants ◽

Murashige And Skoog Medium ◽

Homogeneous Chemical

Cotyledons from developing 6- to 8-week-old embryos of Liatris spicata (L.) Willd. (blazing star) were cultured on Murashige and Skoog medium containing 0, 0.4, 4.4, or 44.4 μm BA or 0, 0.2, 2.2, or 22.2 μm TDZ to induce adventitious shoot formation. The highest percentage of cotyledons forming the most shoots was on medium containing 2.2 μm TDZ. Cotyledon-derived callus cultured on medium containing 4.4 μm BA formed ≈16 times more adventitious shoots than on 2.2 μm TDZ. Adventitious shoots derived from cotyledons or callus produced roots when placed on MS medium containing 5.0 μm IBA. Regenerated plants that flowered in the field appeared homogeneous. Chemical names used: N6-benzyladenine (BA), thidiazuron (TDZ), indole-3-butyric acid (IBA).

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Diallel Analysis of In Vitro Culture Traits in the Genus Lycopersicon

HortScience ◽

10.21273/hortsci.38.1.110 ◽

2003 ◽

Vol 38 (1) ◽

pp. 110-112 ◽

Cited By ~ 4

Author(s):

Guillermo Pratta ◽

Lilians N. Cánepa ◽

Roxana Zorzoli ◽

Liliana A. Picardi

Keyword(s):

Genetic Variability ◽

In Vitro Culture ◽

Opposite Effect ◽

Diallel Analysis ◽

Shoot Formation ◽

Gene Effects ◽

Callus Production ◽

Additive Gene ◽

Productivity Rate

Estimates of genetic variability for in vitro culture traits among the genus Lycopersicon and evaluation of the gene effects involved in callus production and shoot formation were achieved. Five parents including wild and cultivated tomato genotypes and their nonreciprocal 10 possible hybrid combinations were assayed. The callus percentage (C = number of cultures that only produced callus× 100/total number of cultures), the regeneration percentage (R = number of cultures that differentiated into shoots or primordia × 100/total number of cultures) and the productivity rate (PR = total number of shoots/total number of cultures) of each genotype were calculated 45 days after culture initiation. Diallel analysis revealed genetic variability for in vitro culture response. Wild genotypes contributed to a reduction in callus production and an increase in shoot formation while the cultivated genotypes either had an opposite effect or did not modify the expression of culture traits. Hybrids had the lowest callus production and highest shoot formation percentage. Additive gene effects were mainly involved in the expression of C and R, while both additive and nonadditive gene effects were involved in expression of PR.

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Results with the establishment of in vitro culture of Leucojum aestivum

International Journal of Horticultural Science ◽

10.31421/ijhs/13/2/723 ◽

2007 ◽

Vol 13 (2) ◽

Cited By ~ 1

Author(s):

E. Kohut ◽

M. Ördögh ◽

E. Jámbor-Benczúr ◽

Á. Máthé

Keyword(s):

Acetic Acid ◽

Medicinal Plant ◽

In Vitro Culture ◽

In Vitro Cultures ◽

Naphthalene Acetic Acid ◽

Benzyl Adenine ◽

Leucojum Aestivum ◽

Leaves And Roots ◽

Bulbous Plant

Leucojum aestivum is a native, protected ornamental and medicinal plant in Hungary and in Ukraine too. The aim of our work was to establish in vitro cultures of this bulbous plant. Prior to surface sterilisation the old leaves and roots were dissected from the bulbs and they were stored in a refrigerator (2-3°C) for different periods (1 week for the first starting experiment and 5 weeks for the second one). After sterilisation, bulbs, bulb scales and leaves of the bulbs were placed on Murashige and Skoog's (1962) medium with 1 mg/1 benzyl-adenine (BA) and 0,1 mg/1 naphthalene acetic acid (NAA). At the first starting experiment 81,3%, and at the second one 92,3% of the explants turned to be sterile. Bulblets and roots were developed on the explants in the case of using bulb plates together with bulb scales and leaves as inoculua. The best result was achieved after 5 weeks chilling and it was possible to gain little bulbs from the bulb leaves too.

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