DNA Analysis Purdue University Cytometry 脱氧核糖核酸分析普渡大学细胞仪.pptVIP

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DNA Analysis Purdue University Cytometry 脱氧核糖核酸分析普渡大学细胞仪.ppt

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DNA Analysis Purdue University Cytometry 脱氧核糖核酸分析普渡大学细胞仪

DNA1.ppt DNA Probes Key feature of DNA probes is that they are STOICHIOMETRIC - thus number of molecules of probe bound is equivalent to number of molecules of DNA Action of DNA Probes Phenanthridiniums Propidium Iodide Ethidium Bromide Intercalate between bases in DS nucleic acids Excite in the UV or Blue with red emission Bind DS RNA - must be treated with RNAse for DNA measurements to be correct DNA PROBES Action of DNA Probes Benzimides Hoechst 33342 Binds to the A-T rich regions in the small groove of DNA UV excitation with blue emission Red emission if exceed optimal concentration Antibiotics Mithramycin, Chromamycin A3 Bind the G-C rich regions of DNA Action of DNA Probes Acridine Orange - AO Intercalates into the DS DNA Binds to single stranded nucleic acids by stacking on the charged phosphates Blue excitation and green emission for DS DNA Red fluorescence emission for SS DNA Pyronyn Y is a related dye to AO Used for RNA measurement after blocking DNA with non-fluoresceinated DNA probe The Cell Cycle Definitions Terms Ploidy related to the number of chromosomes in a cell Haploid: Number of chromosomes in a gamete (germ cell) is called the HAPLOID number for that particular species Diploid: The number of cells in a somatic cell for a particular species Definitions Terms Hyperdiploid: greater than the normal 2n number of chromosomes Hypodiploid: Less than the normal 2n number of chromosomes DNA Tetraploidy: Containing double the number of chromosomes Definitions Terms DNA Index: The ratio between the mode of the relative DNA content of the test cells (in G0/G1phase) to the mode of the relative DNA content in normal G0/G1 diploid cells Coefficient of Variation - CV: The ratio between the SD of the mode of the G0/G1 cell populations expressed as a percentage. DNA Analysis A typical DNA Histogram Analyzing the DNA Histogram Doublet Discrimination PI Labeled Nuclei Population gating Slide *, * of DNA.ppt ? Purdue University Cytometry Laboratories N+ C2H5 NH2