scholarly journals Determination of content of indole alkaloids in cell biomass of Rauwolfia serpentina Benth. ex Kurz tissue culture

2021 ◽  
Vol 14 (1) ◽  
pp. 73-78
Author(s):  
O. A. Bieda ◽  
I. I. Konvaliuk ◽  
L. P. Mozhylevska ◽  
S. S. Lukashov ◽  
V. A. Kunakh ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Indole Alkaloids ◽  
Natural Plant ◽  
Rauwolfia Serpentina ◽  
Cell Biomass ◽  
Total Alkaloids ◽  
Dry Cell

Cardiovascular diseases are the most common human diseases, hence, the production of cardiological (in particular, anti-arrhythmic) medications from the natural sources is an ever-actual task. Rauwolfia serpentina Benth. is a tropical fruticose plant that is able to produce and concentrate indole alkaloids, especially ajmaline and its derivatives, which are the most effective medications against ventricular arrhythmia with low side effects. Aim of the study. Determination of the qualitative and quantitative content of indole alkaloids in cell biomass of Rauwolfia serpentina tissue culture, obtained by the prolonged in vitro growth. Materials and methods. Object: cell biomass of Rauwolfia serpentina tissue culture (K-27 strain), obtained by methods of long-term cell selection in vitro. Alkaloids content determination: TSQ Vantage LC-MS (ThermoFischer Scientific). Results. 20 indole alkaloids are found in cell biomass of Rauwolfia serpentina tissue culture (K-27 strain). The highest content is registered for ajmaline and its derivatives (0.690 % mass. for ajmaline). The contents of reserpine and yohimbine were found to be as low as 0.009 % and 0.020 %, respectively. Conclusions. It is established that the content of indole alkaloids is higher in K-27 strain in comparison to natural plant and is stable over more than 30 years of its growth. Total alkaloids content was found to be 2.8 % of dry cell biomass, and total ajmaline-type alkaloids content (including ajmaline) was found to be 1.6 % of dry cell biomass. In contrast, the total alkaloid contents in the natural plant material is reported to be in the range of 0.8–1.3 %.

scholarly journals Influence of Indole acetic acid and Tryptophan on production of Vinblastine and Vincristine of Catharanthus roseous callus cells in the accumulation media of In Vitro tissue culture

2010 ◽  
Vol 7 (3) ◽  
pp. 1113-1119
Author(s):  
Baghdad Science Journal
Keyword(s):  
Tissue Culture ◽  
High Performance Liquid Chromatography ◽  
Acetic Acid ◽  
Liquid Chromatography ◽  
In Vitro Culture ◽  
High Performance ◽  
Indole Acetic Acid ◽  
Culture Technique

This study on the plant of Ain –AL Bason Catharanthus roseous showed the ability of callus cells that is produced by In Vitro culture technique and transformed to the accumulated media (MS 40gm/L sucrose ,2gm/L IAA Indole acetic acid , 0.5gm/L Tryptophan) to produce Vinblastine and Vincristine compounds. Extraction, purification and quantitive determination of Vinblastine and Vincristine compounds using High performance liquid chromatography technique (HPLC)were carried out. The results showed that the highest concentration of Vinblastine and Vincristine compounds were ( 4.653,12.5 (ppm /0.5 dry Wight respectively from transformed callus cells from MS 40 gm /L sucrose , 2 gm / L NAA Naphthaline acetic acid .

Characterization of insulin glycation in insulin-secreting cells maintained in tissue culture

1997 ◽  
Vol 152 (1) ◽  
pp. 59-67 ◽  
Author(s):  
Y H A Abdel-Wahab ◽  
F P M O'Harte ◽  
C R Barnett ◽  
P R Flatt
Keyword(s):  
Tissue Culture ◽  
Secretory Activity ◽  
Protein Glycation ◽  
Subsequent Exposure ◽  
Insulin Secreting Cells ◽  
Per Se ◽  
Stimulation Of

Abstract Characteristics of cellular insulin glycation were examined in the pancreatic B-cell line, BRIN-BD11. The extent of insulin glycation increased stepwise during 72 h of culture at 5·6–33·3 mmol/l glucose, attaining levels up to 27%. Glycation of insulin at 33·3 mmol/l glucose was rapid, reaching maximal values within 2 h, and not readily reversible during 2 to 24 h of subsequent exposure to 5·6 mmol/l glucose. Glycated insulin was readily secreted by BRIN-BD11 cells upon active stimulation with glucose and other secretagogues. Cellular insulin glycation was decreased by 66–80% by inhibitors of protein glycation, vitamin C, aminoguanidine or acetylsalicylic acid. Modulation of insulin-secretory activity of BRIN-BD11 cells by co-culture at high glucose with diazoxide, l-alanine or glibenclamide indicated that long-term stimulation of secretion was associated with a decrease in the extent of insulin glycation. Glycation of insulin in vitro was substantially less extensive than in BRIN-BD11 cells, although glucose-6-phosphate and glyceraldehyde-3-phosphate were 1·4- to 2·0-fold more reactive than glucose per se. These observations indicate that insulin is readily glycated and secreted from insulin-secreting cells under hyperglycaemic conditions in culture. Journal of Endocrinology (1997) 152, 59–67

Long-term passage of tachyzoites in tissue culture can attenuate virulence of Neospora caninum in vivo

Parasitology ◽  
2006 ◽  
Vol 133 (4) ◽  
pp. 421-432 ◽  
Author(s):  
P. M. BARTLEY ◽  
S. WRIGHT ◽  
J. SALES ◽  
F. CHIANINI ◽  
D. BUXTON ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Body Weight ◽  
Growth Rates ◽  
Clinical Symptoms ◽  
Neospora Caninum ◽  
High Passage ◽  
Low Passage

To determine whether prolonged in vitro passage would result in attenuation of virulence in vivo, Neospora caninum tachyzoites were passaged for different lengths of time in vitro and compared for their ability to cause disease in mice. Groups of Balb/c mice were inoculated intraperitoneally with 5×106 or 1×107 of low-passage or high-passage N. caninum tachyzoites. The mice were monitored for changes in their demeanour and body weight, and were culled when severe clinical symptoms of murine neosporosis were observed. Mice inoculated with the high-passage parasites survived longer (P<0·05), and showed fewer clinical symptoms of murine neosporosis, compared to the mice receiving the low-passage parasites. The parasite was detected in the brains of inoculated mice using immunohistochemistry and ITS1 PCR. Tissue cysts containing parasites were seen in mice inoculated with both low-passage and high-passage parasites. When the in vitro growth rates of the parasites were compared, the high-passage parasites initially multiplied more rapidly (P<0·001) than the low-passage parasites, suggesting that the high-passage parasites had become more adapted to tissue culture. These results would suggest that it is possible to attenuate the virulence of N. caninum tachyzoites in mice through prolonged in vitro passage.

scholarly journals Determination of two fluoroquinolones and their combinations with hyaluronan effect in in vitro canine cartilage explants

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e6553
Author(s):  
Puntita Siengdee ◽  
Waranee Pradit ◽  
Siriwadee Chomdej ◽  
Korakot Nganvongpanit
Keyword(s):  
Adverse Effects ◽  
Interleukin 1 ◽  
Expression Patterns ◽  
Cartilage Explants ◽  
Beneficial Effects ◽  
Regulated Expression ◽  
Safranin O

Background Previous studies reported the effect of enrofloxacin (Enro) and marbofloxacin (Mar) on cell death and alteration of the key genes involved in catabolic and anabolic processes and demonstrated the beneficial effects of hyaluronan (HA) combined with fluoroquinolones (FQs) on primary canine chondrocytes. This study further determines the effects of these treatments on canine cartilage explants in both normal and interleukin-1 beta (IL-1β)-stimulated conditions. Methods We examined sulfate glycosaminoglycan (s-GAG) release, uronic acid (UA) content, and safranin-O staining, as well as the expression patterns of inflammatory, extracellular matrix (ECM) component and enzymes. Results Enro treatment alone effectively stimulated proteoglycan anabolism by increasing UA content and glycosaminoglycans (GAGs) in normal and pre-IL-1β-stimulated explant, whereas Mar showed opposite results. The combination of HA and FQs increased s-GAG release and UA content in normal explants in addition to effective down-regulated expression of MMP3. HA reduced the adverse effects of Mar by enhancing UA and GAG contents in both normal and pre-IL-1β-explants. Moreover, HA effectively induced HAS1and ACANup-regulation and reduced MMP9, TNF, PTGS2,and NFKB1 expression for a long term. Discussion Our results suggest the direct effects of Enro and Mar may selectively stimulate the conditioned explants to express MMP-codinggenes and promote gene expression involved in matrix production, pro-inflammatory cytokines, and cell degradation in different directions. HA successfully reduced the adverse effects of FQs by enhancing s-GAG and UA contents and down-regulated expression of MMPs.

The use of in vitro methods for plant genetic conservation

1982 ◽  
Vol 11 (2) ◽  
pp. 67-72 ◽  
Author(s):  
C. P. Wilkins ◽  
T. Bengochea ◽  
J. H. Dodds
Keyword(s):  
Tissue Culture ◽  
Genetic Material ◽  
Genetic Conservation ◽  
Fundamental Importance ◽  
Culture Methods ◽  
In Vitro Methods ◽  
Controlled Conditions

The preservation of genetically stable tissue for future propagation is of fundamental importance to plant breeders. In many cases this can be done by storing seed under carefully controlled conditions but there are many plants for which this is not possible or may not be economically feasible. This article reviews current techniques of long-term conservation of plant genetic material by tissue culture methods.

scholarly journals COMPARATIVE STUDIES OF THE ISOZYMES OF LACTIC DEHYDROGENASE IN RABBIT AND MAN

1962 ◽  
Vol 116 (5) ◽  
pp. 797-806 ◽  
Author(s):  
Elliot S. Vesell ◽  
John Philip ◽  
Alexander G. Bearn
Keyword(s):  
Tissue Culture ◽  
Lactic Dehydrogenase ◽  
Isozyme Pattern ◽  
Ldh Activity ◽  
Starch Gel ◽  
Species Specific ◽  
Increased Activity ◽  
Rabbit Tissues

During development of rabbit tissues, characteristic sequential alterations in the LDH isozyme pattern occur, and consist for liver and muscle in loss of the most rapidly migrating anodal bands, and increased activity in the cathodal bands and slower migrating anodal bands. In heart the reverse changes were observed. Comparison of the isozyme patterns observed in various fetal and adult human tissues suggests that these same sequential alterations probably occur. A species-specific isozyme pattern is obtained in long term culture of rabbit, chick, and human cells. The alterations in tissue culture are characterized by a gradual redistribution of total LDH activity in which there is decreased intensity of rapidly migrating anodal bands. These sequential alterations are independent of the organ of origin. The number of bands observed in the starch gel is partly dependent upon the total activities applied. Isozymes may provide a convenient method for determining the species of origin of cell lines in common use and for investigating the effects of various alterations in the in vitro environment on cells grown in tissue culture.

scholarly journals Pengujian Stabilitas Genetik Plantlet Citrumelo Hasil tTCL dari Kultur In Vitro Dengan Menggunakan Teknik Sekuen Berulang

2018 ◽  
Vol 27 (2) ◽  
pp. 165
Author(s):  
Farida Yulianti ◽  
Hidayatul Arisah ◽  
Dita Agisimanto
Keyword(s):  
Tissue Culture ◽  
Genetic Stability ◽  
Simple Sequence Repeat ◽  
Bulk Segregant Analysis ◽  
Cell Layer ◽  
Inter Simple Sequence Repeat ◽  
Transverse Thin Cell Layer ◽  
Simple Sequence

<p>Protokol organogenesis untuk perbanyakan plantlet Citrumelo menggunakan metode transverse thin cell layer (tTCL) batang telah berhasil dikembangkan. Identifikasi stabilitas genetik tanaman hasil kultur jaringan mutlak diperlukan untuk menguji keberadaan off-type. Tujuan penelitian adalah untuk mengetahui potensi primer retrotransposon dan inter simple sequence repeat (ISSR) dalam mendeteksi stabilitas genetik tanaman Citrumelo dari periode kultur yang panjang. Penelitian dilaksanakan pada bulan Juni 2013 sampai dengan Oktober 2015 di Laboratorium Pemuliaan Tanaman, Balai Penelitian Tanaman Jeruk dan Buah Subtropika, Tlekung. Sebanyak empat penanda dengan urutan basa berulang, yaitu retrotransposon dan ISSR digunakan untuk menguji stabilitas genetik plantlet in vitro yang berumur 22 bulan dan untuk mengonfirmasi metode yang dapat diandalkan untuk perbanyakan jeruk Citrumelo yang true-to-type pada masa mendatang. Daun plantlet diseleksi dan diisolasi secara bulk. Amplifikasi dilakukan terhadap DNA dengan sistem bulk segregant analysis (BSA), dan kemudian dipisahkan menggunakan gel agarose. Tanaman in vitro yang sama secara morfologi dapat dibedakan oleh penanda INT-retrotransposon yang mendeteksi adanya kehilangan pita pada grup sampel dengan ukuran 550 bp. Keberadaan retrotransposon dalam genom berlimpah dan aktivasinya diinduksi oleh stres. Kondisi kultur jaringan berpotensi menginduksi aktivasi retrotransposon. Keragaman genetik diperoleh sebesar 2,6%, tetapi masih dapat diterima untuk plantlet yang dihasilkan dari kultur jangka panjang. Plantlet yang digunakan dalam penelitian ini adalah plantlet yang dikulturkan sejak awal tahun 2014 dan telah digunakan untuk mempelajari faktor media dan lingkungan kultur yang efisien pada Citrumelo selama periode 2014–2015. Aktivitas pengkajian variabilitas genetik plantlet yang dihasilkan melalui tTCL batang masih terus dilakukan. Kombinasi protokol dan deteksi berbasis penanda PCR menjadi sarana yang efektif untuk perbanyakan massa benih berkualitas hasil kultur jaringan untuk mendukung progam pemuliaan maupun perbenihan.</p><p>Assessment of genetic stability of long-term cultivation of plantlet derived tTCL Citrumelo using repetitive sequence primers. Regeneration of plantlet from organogenesis of stem transverse thin cell layer (tTCL) was achieved for Citrumelo, a valuable rootstock. Identification of the genetic stability of plant tissue culture is absolutely necessary. The aim of this study was to assess the potential retrotransposons and inter simple sequence repeat (ISSR) primers in detecting the genetic stability of the Citrumelo plantlet derived from tTCL technique. The research was conducted from Juni 2013 until October 2015 in Breeding Laboratory of Indonesian Citrus and Subtropical Fruits Research Institute. A four repetitive based sequences retrotransposon and ISSR marker assays were used to evaluate genetic stability of a group of 22 months old in vitro plantlets and to confirm the most reliable method for true-to-type propagation of Citrumelo. Leaves of plantlets were selected and isolated in bulk. Groups of DNA in bulk segregant analysis (BSA) were amplified and separated using agarose gel. Vitroplants that morphologically similar have been effectively distinguished by a selected primer INT- retrotransposon that detect an deletion band at 550 bp on a line a group of sample. Retrotransposon is abundance through the genome and its activation induced by stress condition. Tissue culture condition was reported potential to induce retrotransposon activation. The genetic variation of 2.6% was acceptable for the culture that produced from long-term. Plantlets used in this study were selected from population induced from early 2014, and employed for studying media as well as environment factors for efficiently organogenesis of citrumelo in period of 2014-2015. However, additional study is on going for evaluating genetic variability from a cycle plantlet production through tTCL of stems. This combination protocol of organogenesis and PCR based markers detection would be powerful tools for mass propagation of high quality seedling derived tissue culture for breeding or cultivation programs.</p>

scholarly journals DEVELOPMENT OF AN ENZYME BIOSENSOR BASED ON pH-SENSITIVE FIELD-EFFECT TRANSISTORS FOR ESTIMATING THE TOTAL CONTENT OF INDOLE ALKALOIDS IN TISSUE CULTURE OF RAUWOLFIA SERPENTINA BENTH. EX KURZ

2021 ◽  
Vol 18 (3) ◽  
pp. 38-50
Author(s):  
V. M. Arkhypova ◽  
О. О. Soldatkin ◽  
L. P. Moghylevska ◽  
І. І. Konvalyuk ◽  
V. А. Kunakh ◽  
...  
Keyword(s):  
Tissue Culture ◽  
Field Effect ◽  
Field Effect Transistors ◽  
Indole Alkaloids ◽  
Total Content ◽  
High Sensitivity ◽  
Ph Sensitive ◽  
Biologically Active ◽  
Rauwolfia Serpentina ◽  
Enzyme Biosensor

A laboratory prototype of an enzyme biosensor based on pH‑sensitive field-effect transistors has been developed to determine the total content of indole alkaloids in Rauwolfia serpentina Benth. Ex Kurz tissue culture. The biosensor was characterized by high sensitivity to th A laboratory prototype of an enzyme biosensor based on pH‑sensitive field effect transistors has been developed to determine the total content of indole alkaloids in Rauwolfia serpentina Benth. Ex Kurz tissue culture. The biosensor was characterized by high sensitivity to the total content of indole alkaloids (minimum limit of determination – 0.5 μg/ml of the total content of indole alkaloids contained in the juice obtained from tissue culture of Rauwolfia serpentina). The linear range of biosensor determination of the analyte was from 2 to 15 μg / ml of the total content of indole alkaloids. Analysis of indole alkaloids using a biosensor is simple and fast and does not require expensive equipment and special sample preparation for analysis, unlike traditional methods. The created biosensor can be further used to control the total content of indole alkaloids in modern biotechnological and pharmaceutical processes for the production of drugs and biologically active additives. e total content of indole alkaloids (minimum limit of determination – 0.5 μg/ml of the total content of indole alkaloids contained in the juice obtained from tissue culture of Rauwolfia serpentina). The linear range of biosensor determination of the analyte was from 2 to 15 μg / ml of the total content of indole alkaloids. Analysis of indole alkaloids using a biosensor is simple and fast and does not require expensive equipment and special sample preparation for analysis, unlike traditional methods. The created biosensor can be further used to control the total content of indole alkaloids in modern biotechnological and pharmaceutical processes for the production of drugs and biologically active additives.  

scholarly journals MOLECULAR-GENETIC ANALYSIS OF PROPAGATED IN VITRO CLONES OF POPULUS ALBA L. AND POPULUS TREMULA L. USING MICROSATELLITE MARKERS

2021 ◽  
Vol 11 (3) ◽  
pp. 16-30
Author(s):  
Tat'yana Grodeckaya ◽  
Oleg Baranov ◽  
Stanislav Rzhevskiy ◽  
Tat'yana Fedulova ◽  
Ekaterina Shabanova ◽  
...  
Keyword(s):  
Molecular Genetic Analysis ◽  
Populus Tremula ◽  
Amplification Coefficient ◽  
Populus Alba ◽  
Time Data ◽  
Electrophoretic Variants ◽  
Reliable Assessment

Use of planting material of forest trees with improved hereditary characteristics is one of the ways to increase the productivity and biological stability of forest stands. It requires taking measures to develop and improve selection base using modern approaches and methods of genetics and biotechnology. A molecular genetics assessment of clone plants of aspen (Populus tremula L.) and white poplar (Populus alba L.) from a long-term in vitro collection (up to 24 years), planted in a greenhouse and field conditions (nursery), was carried out. SSR loci of the PTR series (PTR5, PTR7, PTR8, PTR12, PTR14) were used as DNA markers. Evaluation of clones' ploidy was carried out on the basis of the diagnosis of "loss of heterozygosity" (LOH) effect. Analysis of 5 microsatellite loci of the specimens showed their high intraclonal genotypic stability and homogeneity in vitro and ex vitro. For the first time, data on the results of a comparative determination of ploidy using karyological and microsatellite analysis were presented. Based on the results of the SSR analysis, it can be concluded that the structure of molecular markers is stable among the samples of one clone that are in long-term cultivation. The ratio of the representation (dose) of electrophoretic variants of PCR products serves as an indirect sign of determining ploidy, but for its reliable assessment it is necessary to study the number of loci that are three times larger than the main set of chromosomes. The specimen also requires information on the amplification coefficient of the markers under study. Thus, it is necessary to use both chromosomal and microsatellite analyzes for reliable assessment of intraclonal homogeneity of various specimens, the development of understanding of clone genotypes formation and determination of their ploidy

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