IN VITRO CULTURE OF WHEAT TISSUES. I. CALLUS FORMATION, ORGAN REDIFFERENTIATION AND SINGLE CELL CULTURE

1969 ◽  
Vol 11 (2) ◽  
pp. 294-304 ◽  
Author(s):  
T. Shimada ◽  
T. Sasakuma ◽  
K. Tsunewaki
Keyword(s):  
Cell Culture ◽  
Growth Factors ◽  
Single Cell ◽  
Common Wheat ◽  
Callus Formation ◽  
Casein Hydrolysate ◽  
Callus Growth ◽  
Callus Cell ◽  
Basal Media

Callus induction, organ formation from callus and single callus cell culture have been tried in wheat. Though kinetin showed no effects, supplements of 2,4-D (1~10mg/1) or IAA (50mg/1) to the basal media induced calluses from seedling roots of einkorn, emmer and common wheats, and from stem pieces of common wheat. The best callus growth was obtained when casein hydrolysate (1g/1) or coconut milk (1%) was added to the basal media. Callus growth was also vigorous when 2,4-D (0.5~2.0mg/1) was added. Root formation from callus took place in all kinds of tested media, except those containing no growth factors or supplemented with 2,4-D at high concentrations (1~5mg/1). Shoot formation occurred in six cases and no growth factors were found to be specifically effective on shoot differentiation. Two plants were restored and reached maturity. Calluses of common wheat consisted of eudiploid and aneuploid cells at almost the same frequencies. The great majority of aneuploid cells had 42 ± 3 chromosomes. The restored plants showed normal chromosome constitution. Single callus-cell suspensions were obtained by the liquid culture of seeds in a shaker. A filtrate of the single cell suspension was plated on a solid agar medium, and some colonies were formed. However, plating efficiency was very low and colony growth was slow.

Obtaining and Characterization of Alhagi persarum Boiss. et Buhse Callus Cell Cultures, Isoflavonoid Producers

2020 ◽  
Vol 36 (6) ◽  
pp. 35-48
Author(s):  
D.V. Коchkin ◽  
G.I. Sobolkovа ◽  
А.А. Fоmеnkov ◽  
R.А. Sidorov ◽  
А.М. Nоsоv
Keyword(s):  
Fatty Acids ◽  
Cell Culture ◽  
Liquid Chromatography ◽  
Secondary Metabolites ◽  
Cell Cultures ◽  
Mass Spectrometric ◽  
Mass Spectrometric Detection ◽  
Gas Liquid Chromatography ◽  
Callus Cell

The physiological characteristics of the callus cell cultures of Alhagi persarum Boiss et Buhse, a member of the legume family, widely used in folk medicine, have been studied. It was shown that the source of the explant was an important factor in the initiation of callusogenesis: more intense callusogenesis (almost 100%) was observed for explants from various organs of sterile seedlings, rather than intact plants (less than 30%). As a result, more than 20 lines of morphologically different callus cell cultures were obtained, and the growth parameters for the 5 most intensively growing lines were determined. The composition of fatty acids (FA) of total lipids and secondary metabolites in the most physiologically stable callus line Aр-207 was analyzed. Using capillary gas-liquid chromatography with mass spectrometric detection (GLC-MS), 19 individual C12--C24 FAs were identified, the main fraction of which were palmitic (~ 23%), stearic (~ 22%), linoleic (~ 14%) and α-linolenic (~ 33%) acids. The established atypical ratio of FAs (a simultaneous high content of both saturated FAs and polyunsaturated α-linolenic acid) is possibly due to the adaptation of cells to in vitro growth conditions. Phytochemical analysis of the secondary metabolites was carried out using ultra-performance liquid chromatography with electrospray ionization mass spectrometric detection (UPLC MS). Compounds belonging to different structural groups of isoflavones were found. Aglycones (calycosin, formononetin and afrormosin isomer), glucosides (formononetin glucoside), as well as esters of glucosides (malonylglycosides of calicosin, formononetin, afrormosin isomers, glycitein and genistein) were detected. These secondary metabolites are widespread in plants of the Fabaceae family; however, isoflavones are rare in representatives of the Alhagi genus. The presence of malonylated isoflavone glycosides in Alhagi spp. was shown for the first time. endemic plant species, Alhagi, in vitro cell culture, callus cell culture, isoflavones, fatty acids All studies were carried out using the equipment of the "Experimental Biotechnological Facility" and the "All-Russian Collection of Cell Cultures of Higher Plants" of IРР RAS. This work was supported by the Russian Foundation for Basic Research (RFBR), contract no.18-54-06021 (Az_a), and the Government of the Russian Federation, Megagrant Project no. 075-15-2019-1882.

scholarly journals Pengaruh Komposisi 2,4-D dan BAP Terhadap Pembentukan Kalus Eksplan Pucuk Nilam (Pogostemon Cablin Benth.) secara In Vitro dengan Pemotongan Horizontal dan Vertikal

2021 ◽  
Vol 5 (1) ◽  
pp. 9-20
Author(s):  
Khairan Khairan ◽  
Betty Mauliya Bustam ◽  
Yunita ◽  
Riska Meilinda ◽  
Raudhatul Muna
Keyword(s):  
Acetic Acid ◽  
Callus Formation ◽  
Callus Growth ◽  
Cutting Method ◽  
Highest Weight ◽  
Pogostemon Cablin

This study aims to determine the effect of 2,4-Diclorophenoxy Acetic Acid (2,4-D) and Benzyl Amino Purin (BAP) on the formation of callus of patchouli (Pogostemon cablin Benth.) shoot explants by horizontal and vertical cutting methods. The parameters that observed in this study were the percentage growth of callus, time appearance of callus, weight of callus and the morphology of callus. The results showed that horizontal cutting method was able to induce callus growth with the percentages growth of callus were 18,75%, with the time appearance of callus was at 16 days at P1; P10; P12; P13 dan P14. The highest weight of callus obtained was 0.19 grams at P8. The results also showed that the callus yielded had a yellow and cream color, with a compact and crumb textures. Meanwhile, the vertical cutting method was able to induce callus formation with the percentage growth of callus were 12,5%. The fastest time of callus appearance was obtained in P6 and P8, which was 12 day after planting with the highest weight of callus obtained was 0.05 grams at P12.   The results also showed that vertical cutting method had brown and dark-brown of callus with a compact and crumb textures.

scholarly journals Retention of quiescent hematopoietic cells with high proliferative potential during ex vivo stem cell culture

Blood ◽  
1996 ◽  
Vol 87 (2) ◽  
pp. 545-556 ◽  
Author(s):  
JC Young ◽  
A Varma ◽  
D DiGiusto ◽  
MP Backer
Keyword(s):  
Cell Culture ◽  
Single Cell ◽  
Ex Vivo ◽  
Hematopoietic Cells ◽  
Proliferative Capacity ◽  
Quiescent Cells ◽  
Cell Divisions ◽  
Single Cell Culture ◽  
Bulk Culture

Human CD34+/Thy-1+/Lin- hematopoietic cells purified from bone marrow (BM) or mobilized peripheral blood (MPB) are highly enriched for pluripotent stem cells. Ex vivo expansion of this population is proposed as a means of providing accelerated short-term, as well as long-term, engraftment after myeloablative therapy. Here we demonstrate that primitive quiescent cells are retained in bulk expansion cultures of CD34+/Thy-1+/Lin- cells and that the cell production capacity of the expanded cell product can largely be attributed to cells exhibiting quiescent behavior during culture. CD34+/Thy-1+/Lin- cells from adult BM or MPB were labeled with the fluorescent membrane dye PKH26, followed by in vitro culture of 10(4) cells on a murine stromal layer in the presence of interleukin (IL)-3, IL-6, c-kit ligand (KL), and leukemia inhibitory factor (LIF). With each subsequent cell division, PKH26 fluorescence is reduced by roughly half, which allows tracking of the number of cell divisions. Progenitor cells present after a 2-week expansion period were sorted into CD34+/Lin-/dyebright and CD34+/Lin- /dyedim fractions and then cultured in a 4-week single-cell proliferation assay to characterize the proliferative capacity of each group. Fifty-nine percent of progenitors remaining dyebright after bulk culture (four or fewer cell divisions) were observed to proliferate in single cell culture, and produced an average of 1,780 cells per plated cell. In contrast, only 26% of dyedim (more than four divisions) progenitors were observed to proliferate and displayed a lower average proliferative capacity of 225 cells per plated cell. Similar behaviors were observed after a second consecutive cycle of bulk culture, indicating that quiescent cells with high proliferative capacity existed in culture for at least 4 weeks. Single CD34+/Lin-/dyebright progenitors purified from bulk cultures were observed to produce as many as 1,000 CD34 positive progeny during single cell culture, and these progeny included multilineage colony forming cells. These data demonstrate that among CD34 positive cells recovered after in vitro bulk culture, a higher proliferative capacity correlated with quiescent behavior. The described culture method provides quantitation of the cell producing capacity of individual cells in hematopoietic cell mixtures and may prove useful for predicting engrafting potential in products intended for cellular therapy.

scholarly journals Effect of Surface Sterilization time and Plant Bioregulators for Callus Formation in Hybrid Lilium Cv. Tresor

2017 ◽  
Vol 14 (2) ◽  
pp. 709-713
Author(s):  
Anchal Arpita Gochhayat ◽  
Sashikala Beura ◽  
Enketeswara Subudhi
Keyword(s):  
Callus Formation ◽  
In Vitro Regeneration ◽  
Cut Flowers ◽  
Surface Sterilization ◽  
Basal Media ◽  
Green Callus ◽  
And Control ◽  
Similar Combination ◽  
Effect Of Surface

ABSTRACT: An efficient protocol was standardized for calli mass formation from bulb scale explant of hybrid Lilium Cv. Tresor under in vitro conditions at Biotechnology-cum-Tissue Culture Centre, OUAT, Bhubaneswar. The bulb scale explants were treated with 0.1 % HgCl2 (3 min, 4min, 5min, 6 min, 7min, 8min and 9 min) and control (without treatment) were cultured on MS media, among the treatments, 5 minutes timing resulted in minimum contamination [fungal % (6.67), bacterial % (6.67)] and maximum survival % (83.33%). The best surface sterilization time was further taken into consideration for treatment of explants, sterilization and cultured in the MS Basal media supplemented with BAP (0.5, 1.0 mg/l) in combination with 2,4-D (0.5,1.0,1.5,2.0, and 2.5 mg/l) and 2,4-D (0.5,1.0,1.5,2.0, and 2.5 mg/l) alone along with control. Basal media supplemented with BAP (1.0 mg/l) and 2,4-D (1.50 mg/l) produced maximum callus % (90.00%) and spread, profuse green callus was also recorded in similar combination which opened prospects for developing an indirect means of in vitro regeneration of hybrid Lilium Cv. Tresor there by strengthening the way biotechnology which could be used for improvement and satiate the national and international demands of this cut flowers.

scholarly journals The effect of DL-tryptophan on callus growth and alkaloid-reserpin production of pule pandak [Rauvolfia serpentina (L.) Bentham ex Kurz] in vitro

2005 ◽  
Vol 3 (2) ◽  
pp. 52-56 ◽  
Author(s):  
HENI ARYATI ◽  
ENDANG ANGGARWULAN ◽  
SOLICHATUN SOLICHATUN
Keyword(s):  
Amino Acid ◽  
Quantitative Data ◽  
Callus Formation ◽  
Leaf Explants ◽  
Treatment Phase ◽  
Optimum Concentration ◽  
Callus Growth ◽  
Second Phase ◽  
Rauvolfia Serpentina

The purposes of this research were to study the effect of amino acid DL-tryptophan at various concentrations on culture callus growth and production of alkaloid-reserpine of Rauvolfia serpentina and to determine optimum concentration of DL-tryptophan to yield maximum alkaloid reserpine of the callus. This research was consisted of three phases. First phase was to determine compatible sterilan for R. serpentina leaf explants. Second phase was to initiate/ to induce callus formation from the explants. Third phase was treatment phase to know the influence of DL-tryptophan addition on growth and alkaloid-reserpine production of the callus. Experimental design that used was Completely Randomized Design (CRD). The treatment was concentration of the amino acid DL- tryptophan with three levels concentration three restating for each level. The levels were 0 mg/L, 10 mg/L and 20 mg/L. Obtained data were analyzed quantitatively and qualitatively. Quantitative data were wet weight, dry weight, growth rate and alkaloid content of the callus at each level of concentration. Qualitative data have been measured were compatible sterilant test and callus morphology (color and texture). Quantitative data then have been analyzed by analysis of variance (ANOVA) and continued with DMRT test at level 95%. The result indicated that addition of DL- tryptophan precursor had an effect on the reduction of the callus growth of R. serpentina in media MS in vitro. Addition of DL-tryptophan precursor to produce of alkaloid-reserpine of the callus of R. serpentina in media MS in vitro and optimum concentration of DL-tryptophan precursor that must be added to yield of alkaloid-reserpine could not be determined yet, it caused by the alkaloidreserpine was not detected yet by the thin layer chromatography (TLC).

scholarly journals In Vitro Callus Induction from Leaf Explants of Vanda sp Stimulated by 2,4-D

2017 ◽  
Vol 9 (3) ◽  
pp. 492 ◽  
Author(s):  
Iman Budisantoso ◽  
Nurul Amalia ◽  
Kamsinah Kamsinah
Keyword(s):  
Success Factors ◽  
Callus Formation ◽  
Leaf Explants ◽  
Callus Growth ◽  
Growth Time ◽  
Critical Success ◽  
Randomized Design ◽  
The Difference ◽  
Treatment Concentration

<p class="IsiAbstrakIndo"><span lang="EN-GB">The addition of growth regulator is one of the critical success factors in in vitro cultures. 2,4-D as a plant regulator in media can stimulate the cell division, enlargement of the explants and promotes the formation and growth of callus. The purpose of this study was to determine the time of callus formation and to determine the best concentration of 2.4-D in inducing the growth of callus from leaf explants of </span><em><span lang="EN-GB">Vanda</span></em><span lang="EN-GB"> sp. This research was conducted by experiment with completely randomized design, which consists of six levels of treatment concentration of 2,4-D i.e. 0 ppm; 1 ppm; 1.5 ppm; 2 ppm; 2.5 ppm; and 3 ppm. The parameters observed were the percentage of callus formation and the form of callus from </span><em><span lang="EN-GB">Vanda</span></em><span lang="EN-GB"> sp leaf explants. The results were statistically analyzed by using MINITAB program version 17. Analysis of variance (ANOVA) was performed and the difference between means score/value was separated by F test at p &lt; 0.05. The results showed that 2,4-D treatment give significant effect (F 5,12 = 3,20; p = 0,046 &lt; 0,05) on the callus growth time and its percentage. Application of 2 ppm 2.4-D was the best concentration for accelerating the callus growth time (14.3 days after planting) and increasing the percentage of callus formation (83.3%). Most of callus type were proliferative callus (36.11%) and senescence callus (11.11%). The results of this research are very important to grow the callus from Vanda leaves orchid explant because it is very diffucult to grow.</span></p>

OBTAINING CALLUS CULTURE OF VITEX AGNUS-CASTUS CELLS AS A PRODUCER FOR BIOTECHNOLOGICAL OBTAINING OF PHYTOEXDYSTEROIDS

2021 ◽  
Vol 1 (19) ◽  
pp. 376-377
Author(s):  
S.O. Volodina ◽  
O.V. Topkova
Keyword(s):  
Cell Culture ◽  
Callus Culture ◽  
Callus Cell ◽  
Agnus Castus

The work is devoted to the study of the processes of callusogenesis of Vitex agnus-castus in vitro. The ability of callus cell culture to biosynthesis of ecdysteroids is shown.

scholarly journals Effects of amotosalen treatment on human platelet lysate bioactivity

2019 ◽  
Author(s):  
Christian Christensen ◽  
Sandra Mjoll Jonsdottir-Buch ◽  
Olafur Eysteinn Sigurjonsson
Keyword(s):  
Cell Culture ◽  
Growth Factors ◽  
Blood Donation ◽  
Human Platelet ◽  
Mammalian Cell Culture ◽  
Pathogen Inactivation ◽  
Platelet Concentrates ◽  
Production Methods ◽  
Lineage Differentiation

AbstractBackgroundClinical application of mesenchymal stromal cells (MSCs) usually requires an in vitro expansion step to reach clinically relevant numbers. In vitro cell expansion necessitates supplementation of basal mammalian cell culture medium with growth factors. To avoid using supplements containing animal substances, human platelet lysates (hPL) produced from expired and pathogen inactivated platelet concentrates can be used in place of fetal bovine serum. Due to lack of experience and global diversity in bacterial detection strategies, most transfusion units are currently not pathogen inactivated. As blood banks are the sole source of platelet concentrates for hPL production, it is important to ensure product safety and standardized production methods. To achieve these aims, we assessed the quality of hPL produced from expired platelet concentrates with pathogen inactivation applied after platelet lysis, as well as its ability to support MSC proliferation and tri-lineage differentiation.Methodology/principal findingsBone marrow-derived MSCs (BM-MSCs) were expanded and differentiated using hPL derived from pathogen inactivated platelet lysates (hPL-PIPL), with pathogen inactivation applied after lysis of expired platelets. Results were compared to those using hPL produced from conventional expired pathogen inactivated human platelet concentrates (hPL-PIPC), with pathogen inactivation applied after soon after blood donation. hPL-PIPL treatment had lower concentrations of soluble growth factors and cytokines than hPL-PIPC treatment. When used as supplementation in cell culture, BM-MSCs proliferated at a reduced rate, but more consistently, in hPL-PIPL than in hPL-PIPC. The ability to support tri-lineage differentiation was comparable between lysates.Conclusion/significanceThese results suggest that functional hPL can be produced from expired and untreated platelet lysates by applying pathogen inactivation after platelet lysis. When carried out post-expiration, pathogen inactivation can provide a valuable tool to further standardize global hPL production methods, increase the pool of starting material, and meet the future demand for animal-free supplements in human cell culturing.

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