I am just looking for some clarification into what cells make up a Sub G1 population. I would think that as senescent cells do not lose DNA content they would not be apart of the sub G1 population but rather the G0/G1 population. Is this fair to assume?
sub-G1 is characteristic for low DNA level e.g. apoptotic cells, apoptotic bodies... senescent cells often are charterized as polyploid cells so in my opinion you should connect them with the population G2/M or
In general the sub-G1 "population" is comprised of cell fragments, often as a result of apoptosis, however you may also find aneuploidic cells in here too, it depends on how "sub" the population is. Your assumption is a fair one though, unless senscent cells are more likely to be aneuploidic which I don't think is the case.
I agree with the previous answer. SubG1 cells is characteristic of apoptotic bodies.
sub-G1 is characteristic for low DNA level e.g. apoptotic cells, apoptotic bodies... senescent cells often are charterized as polyploid cells so in my opinion you should connect them with the population G2/M or
I agree with Maciej, in my experience senescent cells are often polyploid cells and are present in the G2/M or higher DNA content populations. Definitely, they do not show a sub-G1 peak. In any case, you can verify the DNA content of the senescent cells you are working with by incubating cells with BrdU for quite long time (e.g. for the duplication time of your cells) and then by double staining them for both BrdU (cycling cells) and DNA content (e.g. 7-AAD).
The sub-G1 peak usually contains apoptotic cells. You could confirm this using Annexin V and PI staining to measure the percentage of apoptotic cells and compare with the percentage of cells in the sub-G1 peak.
Senescent cells often become diploid or polyploidy, whereas dead or dying cells tend to be found in the sub G1 population. Telomere length or telomerase activity would give you an indication of senescence - easier yet though, passage your cells and if they appear to become confluent, but when you count cell number has not really changed from when you split them (e.g. the appearance of confluency is simply because the cells have become bigger), then you know you are dealing with senescent cells. Taking images at the same magnification also provides a good indication of the changing morphology of senescing cells
I Agree with the general asumption of a Sub G1 peak as an expression of apoptotic cells or those that bear a great amount of unrepaired DNA damage. If you plate that cells again you could see a lower plating efficiency and a delay in doubling time.
You may look after DNA damage by using alkaline comet assay that also could allows you to distinguish between damaged and apoptotic cells
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