Plasmid Extraction

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Term Paper Title	Plasmid Extraction
# of Words	397
# of Pages (250 words per page double spaced)	1.59

Plasmid Extraction

Introduction
     Chitobiase, from Vibrio harveyi, is a membrane bound lipoprotein involved in the degradation of chitin.  Chitobiase is similar to and may share a common ancestry to the a-chain of human b-hexos-aminidase.  Chitobiase is encoded by chb.
     In this experiment, a restriction map for restriction enzymes Eco R1, Pst1 and Hind III using Southern hybridization and restriction analysis of pRSG 192.  pRSG 192 is a recombinant plasmid derived from the chb gene and pUC 19, a 2.7kb engineered plasmid which encodes for ampicillin resistance, a portion of the lac operon and a multiple cloning region .  The chb gene exists as a 3.6 kb insert in the mutiple cloning region of pUC 19.
     The major goals of Experiment One will be to isolate pRSG 192 from an overnight culture of      E. coli, amplify a region of the chb gene using PCR, and to map restriction sites within the chb gene using restriction analysis and Southern hybridization.
Methods
Plasmid Isolation
Four microfuge tubes containing cell pellets representing 3.0ml of cells(2 x 1.5ml) from an overnight culture of E. coli were prepared.  The supernatant fluid was discarded an...

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