09-博士研究课生命科学前沿进展多能干细胞-魏炽炬.pptVIP

Induced Pluripotent Stem Cells (iPS) The story of four little genes and a HUGE cellular change 1: Stem Cells Before iPS Embryonic Stem Cells-good source of pluripotent cells, but unethical Somatic cell nuclear transfer-still requires oocytes Stem cells SCNT The basic concept is that the oocyte reprograms the DNA to be “embryonic stem cell-like” Very low efficiency No human stem cell lines have been made from SCNT Not fully reprogramed What is a stem cell? “A cell from which other types of cells can develop.” “An undifferentiated cell that divides to give rise to specialized cells.” “An undifferentiated active somatic cell that undergoes division and gives rise to other stem cells or to cells that differentiate to form specialized cells.” A cell that undergoes asymmetric cell division, giving rise to one stem cell and one partially differentiated cell Types of stem cells In case you missed it… 2: Mouse iPS Techniques and theory Optimization Fibroblasts Are fully differentiated cells Can not become any other cell type Can only divide to make more fibroblasts Contact inhibition If the goal is to get stem cells from normal cells, what would you need to add? Retroviruses Randomly inserts DNA into genome of cells Can make special retroviruses with whatever gene you want Can’t really control how many copies of genes Drug Selection Only turn on a drug resistance gene at stem cell state Do this by using a gene that is only expressed in stem cells Add drug resistance to promoter region of that gene Takes around 16 days for resistance gene to be expressed Drug Selection Four Magic Genes Sox2- Self Renewal Oct4- Differentiation switch Klf4- p53 pathway, Oncogene c-Myc- Global Histone Acetylation, Oncogene Do you really need all 4? Without Oct 3/4 or Klf: no colonies Without Sox2: a few colonies, rough surfaces Without c-Myc: flatter cells, now know actually can do without c-myc-just very low efficiency No Sox2 Are they really stem cells? Somewhere stuck in between Terato