scholarly journals Cryoprotectant treatment tests on three morphologically diverse marine dinoflagellates and the cryopreservation of Breviolum sp. (Symbiodiniaceae)

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Joseph Kanyi Kihika ◽  
Susanna A. Wood ◽  
Lesley Rhodes ◽  
Kirsty F. Smith ◽  
Lucy Thompson ◽  
...  
Keyword(s):  
Genetic Drift ◽  
Rapid Freezing ◽  
Diverse Group ◽  
Limited Success ◽  
The Family ◽  
Agricultural Applications ◽  
Marine Dinoflagellates ◽  
Long Term Preservation ◽  
Cryopreservation Protocol

AbstractDinoflagellates are among the most diverse group of microalgae. Many dinoflagellate species have been isolated and cultured, and these are used for scientific, industrial, pharmaceutical, and agricultural applications. Maintaining cultures is time-consuming, expensive, and there is a risk of contamination or genetic drift. Cryopreservation offers an efficient means for their long-term preservation. Cryopreservation of larger dinoflagellate species is challenging and to date there has been only limited success. In this study, we explored the effect of cryoprotectant agents (CPAs) and freezing methods on three species: Vulcanodinium rugosum, Alexandrium pacificum and Breviolum sp. A total of 12 CPAs were assessed at concentrations between 5 and 15%, as well as in combination with dimethyl sulfoxide (DMSO) and other non-penetrating CPAs. Two freezing techniques were employed: rapid freezing and controlled-rate freezing. Breviolum sp. was successfully cryopreserved using 15% DMSO. Despite exploring different CPAs and optimizing the freezing techniques, we were unable to successfully cryopreserve V. rugosum and A. pacificum. For Breviolum sp. there was higher cell viability (45.4 ± 2.2%) when using the controlled-rate freezing compared to the rapid freezing technique (10.0 ± 2.8%). This optimized cryopreservation protocol will be of benefit for the cryopreservation of other species from the family Symbiodiniaceae.

scholarly journals CRYOPRESERVATION OF TRUE-SEED AND EMBRYO OF MAIZE AND SOYBEAN FOR LONG-TERM STORAGE

2016 ◽  
Vol 2 (2) ◽  
pp. 31
Author(s):  
Enny Sudarmonowati ◽  
I. Fitryatmi ◽  
S. Sadjad
Keyword(s):  
Liquid Nitrogen ◽  
Rapid Freezing ◽  
Soybean Seeds ◽  
Long Term Storage ◽  
Significant Difference ◽  
Long Term Preservation ◽  
Term Storage ◽  
Cryoprotectant Solution ◽  
Excised Embryos

<br />Study on cryopreservation of Indonesian local cultivars and improved  varieties of maize and soybean has never been done. This method may be used for long-term preservation of seeds of maize and soybean. In this study, the method was applied to maize and soybean, Arjuna and Wilis respectively, as a model for preserving germplasm of ortodox seeds. Whole seeds and excised embryos of both varieties were subjected to two methods of cryopreservation, i.e., two-stage cooling and rapid freezing with or without 15% dimethyl sulfoxide (DMSO) as cryoprotectant solution prior to immersion in liquid nitrogen (-196oC). Results indicated that there was no significant difference between the use of DMSO for both species in terms of viability, although pretreatment in DMSO was slightly reduced the percentage of viability of both species. Slow freezing to -30oC prior to immersion in the liquid nitrogen could give as high as 76.67% and 51.67% surviving whole seeds of maize and soybean, respectively. Preserving excised embryos of maize in the liquid nitrogen using either slow or rapid freezing significantly reduced the percentage of viability from 20-76.67% to 5-18.33% (four folds) depending on treatments applied. Results also showed that one day or 15 minutes of immersion of samples in the liquid nitrogen gave rise to similar values of viability of maize and soybean, i.e., 20-60% and 20-51.67%, respectively depending on  treatments applied. These results implied that for long-term storage of maize and soybean seeds as they could survive at the rate of 76.67% and 51.67% respectively, the seed can be treated by prefreezing to -30oC<br />without the presence of DMSO prior to immersion in liquid nitrogen.<br /><br />

scholarly journals Orchid cryopreservation

2014 ◽  
Vol 38 (3) ◽  
pp. 213-229 ◽  
Author(s):  
Wagner Vendrame ◽  
Ricardo Tadeu de Faria ◽  
Mauren Sorace ◽  
Sandra Aparecida Sahyun
Keyword(s):  
Plant Material ◽  
Storage Time ◽  
Natural Habitat ◽  
Orchid Species ◽  
Commercial Use ◽  
The Family ◽  
Relevant Field ◽  
Long Term Preservation ◽  
Intermediate Seeds

Orchids are lush and highly valuable plants due to their diversity and the beauty of their flowers, which increases their commercialization. The family Orchidaceae comprises approximately 35,000 species, distributed among more than 1,000 distinct genera and 100,000 hybrids, totaling approximately 8% to 10% of all flowering plants. With the advance of agriculture and the constant destruction of their natural habitat, orchid species are collected in an indiscriminate manner by collectors and vendors, and this extractive activity threatens many species with extinction, drastically reducing their genetic variability in nature. Therefore, it is essential to seek alternatives that make the preservation of such species feasible using techniques with low maintenance costs that provide greater storage time and that enable good phytosanitary conditions for the plant material for commercial use. Cryopreservation involves the conservation of biological materials at ultra-low temperatures, generally in liquid nitrogen at -196 ºC or in its vapor phase at -150 ºC. This is the only technique currently available for the long-term preservation of the germplasm of plant species that are vegetatively propagated or that have unviable, recalcitrant or intermediate seeds. The objective of this bibliographic review is to report on the importance, methods and application of cryopreservation for orchids. According to the studies reviewed, this is an incipient, developing and relevant field that generates a lot of discussion and requires further research relative to the type of treatment to use for cryopreservation and the methodology to be applied according to the species. The types of methods that are used for cryopreservation and the large variation in the responses of orchids to the cryopreservation methods observed in this study emphasize the need for the development of more appropriate protocols for the preservation of orchids.

scholarly journals Kriopreservasi Tanaman Obat Langka Purwoceng dengan Teknik Enkapsulasi-Vitrifikasi

2016 ◽  
Vol 14 (2) ◽  
pp. 49
Author(s):  
Ika Roostika ◽  
Suci Rahayu ◽  
Novianti Sunarlim
Keyword(s):  
Liquid Nitrogen ◽  
Incubation Period ◽  
Optimization Method ◽  
Endangered Medicinal Plant ◽  
In Vitro Shoots ◽  
Long Term Preservation ◽  
Cryopreservation Protocol ◽  
Cacl2 Solution

<p>Pruatjan (Pimpinella pruatjan Molk.) is an Indonesian endangered medicinal plant, so that it is highly protected. Cryopreservation can be applied to this plant for long-term preservation. The aim of this research was to obtain a method of encapsulation-vitrification by optimizing each step in cryopreservation protocol i.e. preculture, loading, dehydration with and without freezing in liquid nitrogen. The best treatment of each step would be applied in the following step. On preculture experiment, in vitro shoots were planted on the Driver and Kuniyaki (DKW) basal media containing 0.3 M sucrose and incubated for 1, 2, 3, 4, and 5 days. After those incubation period, shoot tips were encapsulated with 2.5% Na-alginate and soaking for 15 minutes in 100 ppm CaCl2 solution before planting. On loading experiment, precultured explants were loaded in DKW basal solution containing 2 M glycerol and 0.4 M sucrose for 0, 30, 60, and 90 minutes. On dehydration experiment, preculturead and loaded explants were dehydrated with PVS2 solution PVS2 (DKW + 30% glycerol + 15% DMSO + 15% ethyleneglicol + 0.4 M sucrose) for 0, 30, 60, 90, and 120 minutes. The parts of them were freezed in liquid nitrogen (-196oC). The result showed that cryopreservation through encapsulation-vitrification technique could be applied on pruatjan. The best preculture treatment was 5 days incubation period. The best loading treatment was 30 minutes. The best dehydration treatment was 90 minutes. The successful level of this research was still low (10%) so that it needs optimization method.</p><p> </p><p><strong>Abstrak</strong></p><p>Purwoceng (Pimpinella pruatjan Molk.) adalah tanaman obat langka asli Indonesia yang hampir punah sehingga harus dilindungi. Kriopreservasi dapat diterapkan pada tanaman ini untuk penyimpanan jangka panjang. Tujuan penelitian adalah untuk memperoleh teknik enkapsulasi-vitrifikasi dengan melakukan optimasi dari tiap-tiap tahapan kriopreservasi yang meliputi perlakuan prakultur, loading, dehidrasi sebelum dan setelah pembekuan dalam nitrogen cair. Perlakuan yang terbaik kemudian diterapkan pada tahapan percobaan berikutnya. Pada perlakuan prakultur, tunas in vitro ditanam pada media Driver dan Kuniyaki (DKW) dengan penambahan sukrosa 0,3 M dengan masa inkubasi 1, 2, 3, 4, dan 5 hari. Setelah itu, pucuk yang berukuran 0,5 cm dienkapsulasi dengan Na-alginat 2,5% (yang mengandung media regenerasi) dalam larutan CaCl2 100 ppm selama 15 menit sebelum penanaman kembali. Pada percobaan loading, terlebih dahulu eksplan diprakultur kemudian direndam dalam larutan DKW + gliserol 2 M + sukrosa 0,4 M dengan durasi rendam selama 0, 30, 60, dan 90 menit. Pada percobaan dehidrasi, eksplan diprakultur dan loading terlebih dahulu, kemudian direndam dalam larutan krioprotektan PVS2 (DKW + gliserol 30% + DMSO 15% + etilen glikol 15% + sukrosa 0,4 M ) selama 0, 30, 60, 90, dan 120 menit. Eksplan tersebut sebagian dibekukan dalam nitrogen cair (-196oC) dan sebagian lainnya tidak dibekukan. Hasil penelitian menunjukkan bahwa kriopreservasi secara enkapsulasi-vitrifikasi berpeluang diterapkan pada tanaman purwoceng. Perlakuan prakultur terbaik adalah 5 hari. Perlakuan loading terbaik adalah 30 menit dan perlakuan dehidrasi terbaik 90 menit. Tingkat keberhasilan ini masih rendah (10%) sehingga diperlukan optimasi metode.</p>

scholarly journals Development of a sperm cryopreservation protocol of Gangetic Mystus (Mystus cavasius)

2017 ◽  
Vol 27 (4) ◽  
pp. 517-529
Author(s):  
MM Islam Islam ◽  
MN Noor ◽  
AA Islam ◽  
MRI Sarder ◽  
MZ Islam ◽  
...  
Keyword(s):  
Dimethyl Sulfoxide ◽  
Sperm Motility ◽  
Genetic Material ◽  
Nacl Solution ◽  
Sperm Cryopreservation ◽  
Endangered Fish ◽  
Long Term Preservation ◽  
Cryopreservation Protocol ◽  
Near Threatened

Cryopreservation is considered as one of the most useful techniques for long-term preservation of genetic material specially sperm of fish. This study focused on the development of a sperm cryopreservation protocol for indigenous near threatened gulsha (Mystus cavasius) and a number of experiments were conducted for the purpose. To collect milt, male gulsha were sacrificed and milt was suspended in extenders. Different concentrations of NaCl were used to evaluate the activation of sperm motility and it decreased as the concentration of the extending media increased, therefore, motility was completely inhibited at 0.8% and 1.2% NaCl solution when sperm suspended in Kurokura-2 and Alsever’s solution, respectively. The toxicity of cryoprotectants to sperm were evaluated using two cryoprotectants, dimethyl sulfoxide (DMSO) and methanol along with the extenders, Alsever’s solution and Kurokura-2 solution. DMSO and methanol with 5% and 10% concentrations produced significantly higher motility during 5 and 10 min incubation and their 15% concentration found toxic to sperm. Alsever’s solution with 10% DMSO produced best equilibration (83.75±2.39%) as well as post-thaw motility (67.5±3.23%) while Kurokura-2 solution with DMSO produced similar equilibration motility (81.25±2.39%) but the post-thaw motility (50.0±6.12%) was significantly much lower than that of Alsever’s solution. Sperm preserved with Alsever’s solution plus DMSO produced highest fertilization, 72.5±7.5% and hatching, 56.8±5.6% while fresh sperm yielded 85.0±5.0% and 74.8±3.6% fertilization and hatching, respectively. The protocols that have been developed can be used for conservation of genetic materials of M. cavasius and other endangered fish species and new generations of them can be propagated using the cryopreserved sperm.Progressive Agriculture 27 (4): 517-529, 2016

scholarly journals Droplet-vitrification methods for apical bud cryopreservation of yacon [Smallanthus sonchifolius (Poepp. and Endl.) H. Rob.]

2021 ◽  
Author(s):  
Stacy Denise Hammond Hammond ◽  
Iva Viehmannova ◽  
Jiri Zamecnik ◽  
Bart Panis ◽  
Milos Faltus
Keyword(s):  
Osmotic Dehydration ◽  
Survival Rates ◽  
Ms Medium ◽  
Smallanthus Sonchifolius ◽  
Apical Bud ◽  
Apical Buds ◽  
Tuberous Roots ◽  
Long Term Preservation ◽  
Cryopreservation Protocol

Abstract This study aimed to develop a cryopreservation protocol for the long-term preservation of yacon [Smallanthus sonchifolius (Poepp. and Endl.)], an Andean crop with high fructooligosaccharide content in its tuberous roots. Initially, the cryopreservation protocol was developed using a yacon clone originated from Ecuador classified as ECU 41. Osmotic dehydration of apical buds (2–3 mm long) was carried out by assessing two plant vitrification solutions, PVS2 (15, 30, and 60 min) at 0°C and PVS3 (30, 45, 60, and 75 min) at 22°C. After cryopreservation, the apical buds were thawed and placed on MS medium ± 0.1 mg l− 1 N6-benzyladenine (BA). The survival rates ranged from 37 to 90% within all treatments, with those subjected to PVS2 and PVS3 for 60 min showing the highest survival rates on MS medium without BA (87 and 90%, respectively). At 12 weeks post cryopreservation, these treatments also provided the highest regrowth rates, both reaching 73% of normally growing (shooting, rooting) plantlets. Survival rates on MS + 0.1 mg l− 1 BA regrowth medium reached up to 90%; however, regrowth into normally rooted plantlets did not exceed 67% post cryopreservation. The optimized protocols were then applied to 4 additional yacon clones originated from Bolivia and Peru, classified as BOL 22, BOL 23, PER 12, and PER 14. This resulted in survival and regeneration rates ranging between 79.7–94.1% and 66.3–75.4% respectively. Our study shows that optimal cryopreservation protocols for the long-term conservation of yacon can be based on both PVS2 and PVS3 vitrification solutions.

scholarly journals CRYOPRESERVATION OF TRUE-SEED AND EMBRYO OF MAIZE AND SOYBEAN FOR LONG-TERM STORAGE

2016 ◽  
Vol 2 (2) ◽  
pp. 31
Author(s):  
Enny Sudarmonowati ◽  
I. Fitryatmi ◽  
S. Sadjad
Keyword(s):  
Liquid Nitrogen ◽  
Rapid Freezing ◽  
Soybean Seeds ◽  
Long Term Storage ◽  
Significant Difference ◽  
Long Term Preservation ◽  
Term Storage ◽  
Cryoprotectant Solution ◽  
Excised Embryos

<br />Study on cryopreservation of Indonesian local cultivars and improved  varieties of maize and soybean has never been done. This method may be used for long-term preservation of seeds of maize and soybean. In this study, the method was applied to maize and soybean, Arjuna and Wilis respectively, as a model for preserving germplasm of ortodox seeds. Whole seeds and excised embryos of both varieties were subjected to two methods of cryopreservation, i.e., two-stage cooling and rapid freezing with or without 15% dimethyl sulfoxide (DMSO) as cryoprotectant solution prior to immersion in liquid nitrogen (-196oC). Results indicated that there was no significant difference between the use of DMSO for both species in terms of viability, although pretreatment in DMSO was slightly reduced the percentage of viability of both species. Slow freezing to -30oC prior to immersion in the liquid nitrogen could give as high as 76.67% and 51.67% surviving whole seeds of maize and soybean, respectively. Preserving excised embryos of maize in the liquid nitrogen using either slow or rapid freezing significantly reduced the percentage of viability from 20-76.67% to 5-18.33% (four folds) depending on treatments applied. Results also showed that one day or 15 minutes of immersion of samples in the liquid nitrogen gave rise to similar values of viability of maize and soybean, i.e., 20-60% and 20-51.67%, respectively depending on  treatments applied. These results implied that for long-term storage of maize and soybean seeds as they could survive at the rate of 76.67% and 51.67% respectively, the seed can be treated by prefreezing to -30oC<br />without the presence of DMSO prior to immersion in liquid nitrogen.<br /><br />

Problems of treatment of primary openangle glaucoma in combination with thrombosis of the central retinal vein

GlaucomaNews ◽  
2020 ◽  
pp. 65-69
Author(s):  
T.E. Lipatkina ◽  
◽  
Е.V. Karlova ◽  
A.V. Zolotarev ◽  
◽  
...  
Keyword(s):  
Macular Edema ◽  
Primary Open Angle Glaucoma ◽  
Antihypertensive Drugs ◽  
Open Angle Glaucoma ◽  
Vascular Wall ◽  
Open Angle ◽  
Retinal Edema ◽  
Long Term Preservation ◽  
Central Retinal Vein

Patients with primary open-angle glaucoma (POAG) and ophthalmic hypertension have an increased likelihood of developing occlusions (thrombosis) of the central retinal vein. Different groups of antihypertensive drugs differ in their mechanism of action and may affect concomitant ocular pathology, in particular, retinal edema, which occurs, for example, in occlusion of the central retinal vein. Used in most patients with glaucoma, prostaglandin analogs can contribute to the long-term preservation of macular edema due to the effect on the permeability of the vascular wall. Preparations of other pharmacological groups, reducing the production of aqueous humor, on the contrary, may contribute to its regression. Therefore, the question of choosing a drug for antihypertensive therapy in patients with primary open-angle glaucoma and concomitant macular edema is relevant and is for further study.

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