Preparation and flow cytometric analysis of metaphase chromosomes of tomato

1991 ◽  
Vol 82 (1) ◽  
pp. 101-111 ◽  
Author(s):  
K. Arumuganathan ◽  
J. P. Slattery ◽  
S. D. Tanksley ◽  
E. D. Earle
Keyword(s):  
Flow Cytometric Analysis ◽  
Metaphase Chromosomes ◽  
Flow Cytometric

Cell synchronization and isolation of metaphase chromosomes from maize (Zea mays L.) root tips for flow cytometric analysis and sorting

Genome ◽  
1996 ◽  
Vol 39 (4) ◽  
pp. 697-703 ◽  
Author(s):  
Jai-Heon Lee ◽  
K. Arumuganathan ◽  
S. M. Kaeppler ◽  
C. M. Papa ◽  
H. F. Kaeppler
Keyword(s):  
Cell Cycle ◽  
Flow Cytometric Analysis ◽  
Maize Root ◽  
Root Tip ◽  
Root Tips ◽  
Chromosome Sorting ◽  
Synthesis Inhibitor ◽  
Metaphase Chromosomes ◽  
Flow Karyotype ◽  
Flow Cytometric

Accumulation of cells containing metaphase chromosomes is an important step in cytological analyses and chromosome sorting procedures. The goal of this research was to optimize treatment parameters to synchronize the cell cycle of maize root tip meristem cells. Levels of hydroxyurea, a DNA synthesis inhibitor, were assessed for their utility in accumulating cells at the G1 phase of the cell cycle. Trifluralin, amiprophos-methyl, and colchicine were used to accumulate cells containing metaphase chromosomes upon release from hydroxyurea inhibition. Optimal mitotic indices were achieved by treating seedlings with 5 mM hydroxyurea for 18 h, incubating for 1 h without chemical treatment to release the hydroxyurea block, and then treating emerging roots with 1 μM trifluralin for 4 h. The mitotic index of synchronized maize root tips was over 70%. Uniformity of synchronization depended upon selection of seeds with emerging radicles that were similar in length at the time of treatment. Suspensions of intact chromosomes were prepared by a simple slicing procedure. The chromosome preparations were found to be suitable for flow cytometric characterization and sorting. Chromosome peaks of the observed flow karyotype resembled the predicted flow karyotype calculated on the basis of maize chromosome size. Key words : flow karyotype, hydroxyurea, plant chromosome sorting, trifluralin.

scholarly journals Isolated metaphase chromosomes. II. Proteins of Chinese hamster chromosomes.

1979 ◽  
Vol 27 (1) ◽  
pp. 454-457 ◽  
Author(s):  
W Wray ◽  
V P Wray
Keyword(s):  
Flow Cytometric Analysis ◽  
Chinese Hamster ◽  
High Capacity ◽  
Microscopic Analysis ◽  
Metaphase Chromosomes ◽  
Chromosomal Proteins ◽  
Quantitative Evidence ◽  
Flow Cytometric ◽  
Velocity Gradients

The proteins on metaphase chromosomes theoretically may be distributed ubiquitously throughout the karyotype, may be present uniquely on individual chromosomes or classes of chromosomes, or may exist in any combination of the above. Separation of chromosomes according to size using sucrose velocity gradients in high capacity zonal centrifuge rotors allows sufficient fractionation of the genome to indicate the distribution of proteins within the karyotype. Flow cytometric analysis and direct microscopic analysis were used to evaluate qualitatively the types of chromosomes present in the fractions obtained. This report is the first quantitative evidence that some of the chromosomal proteins are not distributed ubiquitously on all of the chromosomes of the karyotype.

Root tip cell cycle synchronization and metaphase-chromosome isolation suitable for flow sorting in common wheat (Triticum aestivum L.)

Genome ◽  
1997 ◽  
Vol 40 (5) ◽  
pp. 633-638 ◽  
Author(s):  
Jai-Heon Lee ◽  
K. Arumuganathan ◽  
Y. Yen ◽  
S. Kaeppler ◽  
H. Kaeppler ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Common Wheat ◽  
Univariate Analysis ◽  
Flow Cytometric Analysis ◽  
Root Tip ◽  
Bivariate Analysis ◽  
Root Tips ◽  
Metaphase Chromosomes ◽  
Cell Cycle Synchronization ◽  
Flow Cytometric

An efficient procedure for cell-cycle synchronization in meristematic root tips was achieved in common wheat. Treatment parameters for synchronizing the cell cycle of root tip meristem cells, such as time-course and applied concentrations of various chemicals, were systematically tested and optimized by flow cytometric analysis of isolated nuclei. High mitotic indices (69.5% in the root tip meristematic area) were routinely obtained by treating germinating seeds with 1.25 mM hydroxyurea for 16 h, followed by incubation in a hydroxyurea-free solution for 2 h, and treatment with 1 μM trifluralin for 4 h. Uniform seed germination prior to treatment is very important for achieving consistently high metaphase indices in the root tips. Large numbers of metaphase chromosomes, suitable for flow cytometric analysis and sorting, were isolated from synchronized root tip cells. Flow sorted wheat chromosomes, via univariate and bivariate analysis, showed four major chromosome peaks. Each discrete peak may represent wheat chromosome types with similar DNA content. Bivariate flow karyotyping based on AT and GC content did not improve the separation of wheat chromosomes.Key words: flow cytometry, trifluralin, hydroxyurea, univariate analysis, bivariate analysis.

Apoptotic Effects of Melittin on 4T1 Breast Cancer Cell Line is associated with Up Regulation of Mfn1 and Drp1 mRNA Expression

2020 ◽  
Vol 20 (7) ◽  
pp. 790-799 ◽  
Author(s):  
Farnaz D. Moghaddam ◽  
Pejman Mortazavi ◽  
Somayeh Hamedi ◽  
Mohammad Nabiuni ◽  
Nasim H. Roodbari
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Hemolytic Activity ◽  
Breast Cancer Cells ◽  
Flow Cytometric Analysis ◽  
Control Group ◽  
Flow Cytometric ◽  
4T1 Cells ◽  
4T1 Breast Cancer ◽  
Apoptotic Effects

Background and Purpose: Melittin, as the main ingredient of honeybee venom, that has shown anticancer properties. The present study aimed at investigating the cytotoxic impacts of melittin on 4T1 breast cancer cells. Methods: Hemolytic activity of different concentrations (0.125, 0.25, 0.5, 1, 2, 4, 8μg/ml) of melittin was assayed and then cytotoxicity of selected concentrations of melittin (2, 4, 8, 16, 32, and 64μg/ml), 2 and 4μg/ml of cisplatin and 0.513, 0.295 and 0.123μg/ml of doxorubicin was evaluated on 4T1 cells using MTT assay. We used Morphological evaluation and flow cytometric analysis was used. Real time PCR was also used to determine mRNA expression of Mfn1 and Drp1 genes. Results: All compounds showed anti-proliferative effects on the tumor cell line with different potencies. Melittin had higher cytotoxicity against 4T1 breast cancer cells (IC50= 32μg/ml-72h) and higher hemolytic activity (HD50= 1μg/ml), as compared to cisplatin and doxorubicin. Mellitin at 16 and 32μg/ml showed apoptotic effects on 4T1 cells according to the flow cytometric analysis. The Real time PCR analysis of Drp1 and Mfn1 expression in cells treated with 16μg/ml of melittin revealed an up-regulation in Drp1 and Mfn1 genes mRNA expression in comparison with control group. Treatment with 32μg/ml of melittin was also associated with a rise in mRNA expression of Drp1 and Mfn1 as compared to the control group. Conclusion: The results of this study showed that melittin has anticancer effects on 4T1 cell lines in a dose and time dependent manner and can be a good candidate for further research on breast cancer treatment.

scholarly journals Flow cytometric analysis of T lymphocytes and cytokines in aqueous humor of patients with varicella zoster virus-mediated acute retinal necrosis

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Hao Kang ◽  
Yunbo Wei ◽  
Ming Liu ◽  
Di Yu ◽  
Yong Tao
Keyword(s):  
T Lymphocytes ◽  
Varicella Zoster Virus ◽  
Aqueous Humor ◽  
T Lymphocyte ◽  
Lymphocyte Subsets ◽  
Flow Cytometric Analysis ◽  
T Lymphocyte Subsets ◽  
Varicella Zoster ◽  
Flow Cytometric ◽  
Cd8 T Lymphocytes

Abstract Background The purpose of this study is to investigate the aqueous humor (AH) T lymphocyte subsets and cytokines of acute retinal necrosis (ARN) to elucidate the immunologic inflammatory features of this disorder. Methods Three patients with ARN infected with varicella zoster virus (VZV) who underwent multiple intravitreal injections of ganciclovir were enrolled in this study. The control group consisted of four non-infectious patients with acute anterior uveitis (AAU). Flow cytometric analysis was performed on the lymphocyte subsets from the AH and peripheral blood (PB) samples during the active phase of intraocular inflammation. Five inflammatory cytokines were measured in each AH sample and various clinical characteristics were also assessed. Results VZV deoxyribonucleic acid (DNA) was detected by real-time polymerase chain reaction (PCR) in AH from all the ARN patients, who showed higher CD8+ T lymphocytes population in AH than the AAU patients (p = 0.006). CD4/CD8 ratios of T lymphocytes and the percentage of CD8 + CD25+ T lymphocytes in AH were significantly lower in ARN than in AAU (p = 0.006; p = 0.012). In the ARN patients, the percentages of CD4+ and CD8+ T lymphocytes in AH were higher than those found in PB. The percentage of CD4 + CD25+ T lymphocytes in AH was significantly higher than the proportion in PB in the AAU patients (p = 0.001). Immunoregulatory cytokine Interleukin-10 in AH was significantly elevated in the ARN patients in comparison with the case of the AAU patients (p = 0.036). In ARN, the copy number of VZV DNA in AH positively correlated with the percentage of CD8+ T lymphocytes in AH and negatively correlated with the CD4/CD8 ratio in AH during the course of disease treatment (p = 0.009, r = 0.92; p = 0.039, r = − 0.834). Conclusion The ARN patients caused by VZV had different intraocular T lymphocyte subsets and cytokines profile than those of the non-infectious patients. High percentages of CD8+ T lymphocytes and low CD4/CD8 T cell ratios may be a potential biomarker for diagnosis of viral-infectious uveitis. T lymphocytes examination at the inflammatory sites has the potential to become a useful research tool for differentiating viral and non-viral uveitis.

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