NME variant species expression and suppression
A variant, biological technology, applied in the direction of biochemical equipment and methods, microorganisms, drug combinations, etc., can solve problems such as not understanding the functional relationship of NM23
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Embodiment 1
[0226] Example 1 describes an experiment in which NM23-S120G of BGO1v human embryonic stem cells (which have been renatured and purified to exist mainly as dimers) on a coating of anti-MUC1* monoclonal antibody MN-C3 or cultured in bFGF on mouse fibroblast feeder cells. Western blot of the resulting cells showed that NME7 was highly expressed in stem cells that had been cultured in NM23-S120G dimers ( figure 1 , part I.C—lane 1), but only weakly expressed in stem cells cultured in bFGF (lane 2). We have shown that stem cells cultured in NM23 dimers revert to a naïve state (Smagghe et al., 2013) compared to the primed state of human stem cells cultured in bFGF due to both commercially available stem cell lines. , the naive state is a less mature state. The fact that NME7 is more expressed in naive state stem cells than in prime state stem cells is consistent with the fact that NME7 is preferentially expressed at very early stages of embryogenesis rather than later in embryoni...
Embodiment
[0253] Example 1 To determine whether human stem cells express NME6 or NME7 in addition to NME1 (H1) and NME2 (H2), we performed Western blot analysis on lysates and supernatants from various human stem cell lines. 4 ng / mL in a) only the dimeric form of NM23-S120G on cell culture plates coated with anti-MUC1* monoclonal antibody MN-C3, or b) on mouse feeder cells (MEF) Human embryonic stem cell line BGO1v cells were cultured under bFGF. After 3 days in culture, stem cells were harvested and lysed, then analyzed by Western blot using antibodies to detect the presence of NME1, NME6 and NME7. For comparison, the same analysis was performed in parallel on T47DMUC1*-positive breast cancer cells. As a control, recombinant NM23-H1 wild-type (NM23-wt) protein was loaded on the gel and also probed with antibodies recognizing 3 different NMEs. Note that the gel is a denaturing gel such that the apparent molecular weights of the NM23-S120G dimer and wild-type hexamer are expressed as t...
Embodiment 2
[0259] Example 2. Western blot analysis of the human stem cell line BGO1v and HES-3 cells showed that a purportedly specific NME antibody against NME7 recognized NME7 as well as another species with an apparent molecular weight of ~22-25 kDa ( figure 2 ).
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