scholarly journals Effect of Cryopreservation Method Supported With Biochemical Analyses in the Axillary Bud of Jewel Orchid, Ludisia Discolor

2021 ◽  
Author(s):  
Hazirah Burkhan ◽  
Kirutika Selva Rajan ◽  
Suganthi Appalasamy ◽  
Ranjetta Poobathy ◽  
Bee Lynn Chew ◽  
...  
Keyword(s):  
Abiotic Stresses ◽  
Axillary Buds ◽  
Oxygen Species ◽  
Poor Survival ◽  
Varied Response ◽  
Biochemical Analyses ◽  
Jewel Orchid ◽  
Cryopreservation Method ◽  
Recovery Medium

Abstract This study was conducted to investigate the potential of conserving an endangered terrestrial jewel orchid Ludisia discolor using in vitro grown axillary buds. Excised segments of axillary buds (4 - 5 mm in length) were precultured on a modified Murashige and Skoog (MS) medium supplemented with 0.2 M sucrose for 24 h and osmoprotected in a loading solution for 20 min. Then, axillary buds were dehydrated in PVS2 solution for 10 min at 0°C and incubated in liquid nitrogen for 1 h. Subsequently, axillary buds were rewarmed rapidly by dilution solution and transferred to a growth recovery medium supplemented with 0.05 µM melatonin under in vitro conditions that led to an improved survival chance (16.67%) for cryopreserved L. discolor. The abiotic stresses and the overproduction of reactive oxygen species (ROS) during cryopreservation stages may contribute to cryoinjuries and poor survival. The varied response towards stress was detected with significantly increased values recorded at certain cryopreservation stages, including proline activity at the dehydration stage (5.51 µmol/g), catalase at the preculture (85.64 U/g) and dehydration (70.87 U/g) stages, peroxidase at the rewarming stage (565.37 U/g) and ascorbate peroxidase during the loading stage (12.19 U/g). Hence, this first attempt to cryopreserved L. discolor indicates that future experimental designs could include exogenous antioxidants and different vitrification solutions to improve survival and regeneration.

scholarly journals Moderate abiotic stresses increase rhizome growth and outgrowth of axillary buds in Alstroemeria cultured in vitro

2012 ◽  
Vol 110 (3) ◽  
pp. 395-400 ◽  
Author(s):  
Paweena Pumisutapon ◽  
Richard G. F. Visser ◽  
Geert-Jan de Klerk
Keyword(s):  
Abiotic Stresses ◽  
Axillary Buds ◽  
Rhizome Growth

Fine cryopreservation method of porcine blastocysts produced by in vitro fertilization

2014 ◽  
Author(s):  
Sung-Hun Min ◽  
Jin-Woo Kim ◽  
Yong-Hee Lee ◽  
Jae-Hyun Ahn ◽  
Geon-Yeop Do ◽  
...  
Keyword(s):  
In Vitro Fertilization ◽  
Vitro Fertilization ◽  
Cryopreservation Method

scholarly journals Gold nanoparticles affect the cryopreservation efficiency of in vitro-derived shoot tips of bleeding heart

2021 ◽  
Author(s):  
Dariusz Kulus ◽  
Alicja Tymoszuk
Keyword(s):  
Gold Nanoparticles ◽  
Alginate Bead ◽  
Stability Index ◽  
Shoot Tips ◽  
Start Codon ◽  
Lamprocapnos Spectabilis ◽  
Colloid Dispersion ◽  
Recovery Medium ◽  
Alginate Matrix

AbstractThe popularity of nanoparticles (NPs) is continuously increasing. To date, however, there has been little research on the application of NPs in plant cryopreservation, i.e. storage of tissues in liquid nitrogen (LN). The aim of this study is to analyze the effect and evaluate the usefulness of gold nanoparticles (AuNPs) in regard to cryobiology studies. In vitro-derived shoot tips of Lamprocapnos spectabilis ‘Valentine’ were cryopreserved with the encapsulation-vitrification protocol. Gold nanoparticles (at 10–30 ppm concentration; 13 nm in size) were added either into the preculture medium; to the protective bead matrix during encapsulation; or to the recovery medium after rewarming of samples. The control plants were produced from cryopreserved explants non-treated with nanoparticles or treated with colloid dispersion medium without NPs. A non-LN-treated standard was also considered. The influence of AuNPs on the cryopreservation efficiency was determined by evaluating the recovery rate of explants and their morphogenic response; the membrane stability index (MSI); the concentration of pigments in shoots; and the antioxidant enzymes activity. The genetic stability of the plant material was evaluated using Start Codon Targeted Polymorphism (SCoT) markers. It was found that 10 ppm of AuNPs added into the alginate bead matrix improved the recovery level of LN-derived shoot tips (70.0%) compared to the non-NPs-treated cryopreserved control (50.5%). On the other hand, the presence of nanoparticles in the recovery medium had a deleterious effect on the survival of explants. AuNPs usually had no impact on the MSI (73.9–85.9%), except for those added into the recovery medium at the concentration of 30 ppm (decline to 55.8%). All LN-derived shoots were shorter and contained less chlorophyll and carotenoids than the untreated standard. Moreover, the application of AuNPs affected the enzymatic activity in L. spectabilis. Minor genetic variation was found in 8.6% of plants if AuNPs were added either into the preculture medium (at 10 and 20 ppm) or to the alginate matrix (at 30 ppm). In conclusion, AuNPs added at a lower concentration (10 ppm) into the protective bead matrix can significantly improve the cryopreservation efficiency in L. spectabilis with no alternation in the DNA sequence.

scholarly journals Apoptosis-like cell death upon kinetoplastid induction by compounds isolated from the brown algae Dictyota spiralis

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Olfa Chiboub ◽  
Ines Sifaoui ◽  
Manef Abderrabba ◽  
Mondher Mejri ◽  
José J. Fernández ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Cell Death ◽  
Mitochondrial Membrane ◽  
Brown Seaweed ◽  
Cell Permeability ◽  
Oxygen Species ◽  
Potential Drug ◽  
Concentration Cell ◽  
Drug Candidates

Abstract Background The in vitro activity of the brown seaweed Dictyota spiralis against both Leishmania amazonensis and Trypanosoma cruzi was evaluated in a previous study. Processing by bio-guided fractionation resulted in the isolation of three active compounds, classified as diterpenes. In the present study, we performed several assays to detect clinical features associated to cell death in L. amazonensis and T. cruzi with the aim to elucidate the mechanism of action of these compounds on parasitic cells. Methods The aims of the experiments were to detect and evaluate specific events involved in apoptosis-like cell death in the kinetoplastid, including DNA condensation, accumulation of reactive oxygen species and changes in ATP concentration, cell permeability and mitochondrial membrane potential, respectively, in treated cells. Results The results demonstrated that the three isolated diterpenes could inhibit the tested parasites by inducing an apoptosis-like cell death. Conclusions These results encourage further investigation on the isolated compounds as potential drug candidates against both L. amazonensis and T. cruzi. Graphic abstract

scholarly journals Efficient Cryopreservation of Populus tremula by In Vitro-Grown Axillary Buds and Genetic Stability of Recovered Plants

Plants ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 77
Author(s):  
Elena O. Vidyagina ◽  
Nikolay N. Kharchenko ◽  
Konstantin A. Shestibratov
Keyword(s):  
Survival Rate ◽  
Axillary Buds ◽  
Long Term Storage ◽  
Average Survival ◽  
Transgenic Lines ◽  
Ssr Loci ◽  
One Step ◽  
The One

Axillary buds of in vitro microshoots were successfully frozen at –196 °C by the one-step freezing method using the protective vitrification solution 2 (PVS2). Microshoots were taken from 11 transgenic lines and three wild type lines. Influence of different explant pretreatments were analyzed from the point of their influence towards recovery after cryopreservation. It was found out that the use of axillary buds as explants after removal of the apical one increases recovery on average by 8%. The cultivation on growth medium of higher density insignificantly raises the regenerants survival rate. Pretreatment of the osmotic fluid (OF) shows the greatest influence on the survival rate. It leads to the increase in survival rate by 20%. The cryopreservation technology providing regenerants average survival rate of 83% was developed. It was based on the experimental results obtained with explant pretreatment. Incubation time in liquid nitrogen did not affect the explants survival rate after thawing. After six months cryostorage of samples their genetic variability was analyzed. Six variable simple sequence repeat (SSR) loci were used to analyze genotype variability after the freezing-thawing procedure. The microsatellite analysis showed the genetic status identity of plants after cryopreservation and of the original genotypes. The presence of the recombinant gene in the transgenic lines after cryostorage were confirmed so as the interclonal variation in the growth rate under greenhouse conditions. The developed technique is recommended for long-term storage of various breeding and genetically modified lines of aspen plants, as it provides a high percentage of explants survival with no changes in genotype.

scholarly journals The Effects of Bioactive Compounds from Blueberry and Blackcurrant Powder on Oat Bran Pastes: Enhancing In Vitro Antioxidant Activity and Reducing Reactive Oxygen Species in Lipopolysaccharide-Stimulated Raw264.7 Macrophages

Antioxidants ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 388
Author(s):  
Xiao Dan Hui ◽  
Gang Wu ◽  
Duo Han ◽  
Xi Gong ◽  
Xi Yang Wu ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Antioxidant Activity ◽  
Phenolic Compounds ◽  
Antioxidant Capacity ◽  
Bioactive Compounds ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Raw264.7 Macrophages ◽  
Oat Bran

In this study, blueberry and blackcurrant powder were chosen as the phenolic-rich enrichments for oat bran. A Rapid Visco Analyser was used to form blueberry and blackcurrant enriched oat pastes. An in vitro digestion process evaluated the changes of phenolic compounds and the in vitro antioxidant potential of extracts of pastes. The anthocyanidin profiles in the extracts were characterised by the pH differential method. The results showed that blueberry and blackcurrant powder significantly increased the content of phenolic compounds and the in vitro antioxidant capacity of pastes, while the total flavonoid content decreased after digestion compared to the undigested samples. Strong correlations between these bioactive compounds and antioxidant values were observed. Lipopolysaccharide-stimulated RAW264.7 macrophages were used to investigate the intracellular antioxidant activity of the extracts from the digested oat bran paste with 25% enrichment of blueberry or blackcurrant powder. The results indicated that the extracts of digested pastes prevented the macrophages from experiencing lipopolysaccharide (LPS)-stimulated intracellular reactive oxygen species accumulation, mainly by the Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2) signalling pathway. These findings suggest that the bioactive ingredients from blueberry and blackcurrant powder enhanced the in vitro and intracellular antioxidant capacity of oat bran pastes, and these enriched pastes have the potential to be utilised in the development of the functional foods.

scholarly journals Heterodimer Formation of the Homodimeric ABC Transporter OpuA

2021 ◽  
Vol 22 (11) ◽  
pp. 5912
Author(s):  
Patricia Alvarez-Sieiro ◽  
Hendrik R. Sikkema ◽  
Bert Poolman
Keyword(s):  
Abc Transporter ◽  
Conformational Dynamics ◽  
Host Organism ◽  
Affinity Tag ◽  
Practical Applications ◽  
Fundamental Research ◽  
Protein Multimerization ◽  
Biochemical Analyses

Many proteins have a multimeric structure and are composed of two or more identical subunits. While this can be advantageous for the host organism, it can be a challenge when targeting specific residues in biochemical analyses. In vitro splitting and re-dimerization to circumvent this problem is a tedious process that requires stable proteins. We present an in vivo approach to transform homodimeric proteins into apparent heterodimers, which then can be purified using two-step affinity-tag purification. This opens the door to both practical applications such as smFRET to probe the conformational dynamics of homooligomeric proteins and fundamental research into the mechanism of protein multimerization, which is largely unexplored for membrane proteins. We show that expression conditions are key for the formation of heterodimers and that the order of the differential purification and reconstitution of the protein into nanodiscs is important for a functional ABC-transporter complex.

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