biology 302 lab 2

biology 302 lab 2 - Introduction The major question in this...

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Introduction The major question in this experiment was what PMA or DMSO HL-60 cells would produce when undergoing reverse transcriptase and PCR. We performed this experiment to see cell differentiation on a transcriptase level. We are studying this because HL-60 cells are multipotent cells which means they can differentiate into different cell types. In this study we are observing how the PMA and DMSO cells induce a corresponding protein. The proteins produced in this experiment are Beta- Actin and MMP-9. It is believed that Beta-Actin is produced by monocytes and granulocytes which correspond to both PMA and DMSO HL-60 cells since PMA is known to produce monocytes while DMSO is known to produce granulocytes. It is also believed that MMP-9 is only produced by monocytes which correspond only to the PMA HL-60 cells. We intend to prove that the PMA HL-60 cells will produce both types of protein while DMSO will only produce Beta-Actin. Materials and Methods Before beginning this experiment use the dye test to make sure the cells are below 3*10^6. First, we obtained 5 mL of untreated PMA HL-60 cells. We centrifuged the cells for 2 minutes at 12000 rpms to pellet the cells. We then removed all supernatant from the tube. Next, we added 300 microliters of RNA Protect Cell Reagent to the cell pellet. We re-suspended the cells by vortexing them for five seconds. Next we added 350 microliters of buffer RLT to disrupt and homogenize the cells. After disrupting and homogenizing the cells, we vortexed the cells for 5 seconds.
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We then transferred the cell lysate into a Purple QIA shredder spin column. We
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biology 302 lab 2 - Introduction The major question in this...

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