flow cytometry with a partec cell counter analyzer cca-ii  (Partec)

Journal:

Article Title: Analysis of Drosophila Species Genome Size and Satellite DNA Content Reveals Significant Differences Among Strains as Well as Between Species

doi: 10.1534/genetics.107.075069

Figure Lengend Snippet: Drosophila polyploid follicle cells underreplicate satellite DNA repeats. Proliferating follicle cells duplicate their entire genomes and cycle from 2C to 4C and after mitotic division back to 2C (A). 2C cells enter their polyploid state by replicating their euchromatic sequences and replicate little or no centric/pericentric satellite repeat sequences (B). Consequently, 4c-p cells have less 4C DNA content, and a second and third round of polyploid S-phases produce 16C cells with vastly underreplicated satellite DNA. Flow cytometry histograms of follicle cell nuclei from (C) D. melanogaster, (D) D. grimshawi, (E) D. immigrans, and (F) D. virilis are shown by illustrating the four major 2C, 4C, 8C, and 16C ploidy peaks where the x-axis represents arbitrary fluorescent units and the y-axis is the number of nuclei. Note that the 4C peak can be resolved into two peaks (see insets in C and F), where the 4C peak from mitotic proliferating cells has more DNA content than the 4C-p peak. This is because follicle cells undergoing polyploidization fail to replicate the centric and pericentric satellite repeats and thus have less DNA than mitotic 4C cells, as described in A. In larger genomes such as (D) D. grimshawi, (E) D. immigrans and (F) D. virilis, the extent of underreplication can be seen by a dramatic shift of all polyploid peaks to the left. The most extreme example is seen in (F) D. virilis where the 8C peak nearly overlaps the normal mitotic cell 4C peak (see inset), suggesting that about half of the genome fails to replicate. This is consistent with measurements of ∼48% heterochromatin content in D. virilis (see Table 5). We observed underreplication in all 91 strains from all 38 species that we examined.

Article Snippet: Samples were analyzed on a PARTEC CCA-II flow cytometry machine (PARTEC).

Techniques: Flow Cytometry

Journal:

Article Title: Analysis of Drosophila Species Genome Size and Satellite DNA Content Reveals Significant Differences Among Strains as Well as Between Species

doi: 10.1534/genetics.107.075069

Figure Lengend Snippet: Larger genomes have greater underreplication. The percentage of underreplication (y-axis) was calculated on the basis of 16C/2C values (see materials and methods) and is shown plotted against haploid genome size in megabases, as determined by PI (A) and DAPI (B) flow cytometry. A trend line was added to show that, as genomes become larger, a greater fraction of the total DNA content is underreplicated. Note that the same trend is observed regardless of the dye used. Two-tailed P-values were calculated as in Figure 3.

Article Snippet: Samples were analyzed on a PARTEC CCA-II flow cytometry machine (PARTEC).

Techniques: Flow Cytometry, Two Tailed Test