Molecular Diagnostics Chapter 9 Exam with verified solutions

Molecular Diagnostics Chapter 9 Exam with verified solutions Maxam-Gilbert Sequencing - ANSWERDiffering concentrations of salt are used in four different tubes - A, T, C, G - Usually DMS (dimethylsulphate), FA (formic acid), H (hydrazine), and H+S (hydrazine + salt) which cuts nucleic acids at certain nucleotides A, GA, TC, C-- read on a gel from bottom up, Sanger sequencing was adopted in labs instead since it can sequence longer fragments and doesn't use as toxic of chemicals dideoxy chain termination - ANSWERSanger sequencing, the early method of sequencing DNA Sanger sequencing - ANSWERDideoxynucleotides halt DNA polymerization at each base since they lack the hydroxyl on the 3' ribose carbon and cannot form a phosphodiester bond, the template along with DNTP's and PCR MM is added to four different tubes with a different ddNTP in each, generating sequences of various lengths that encompass the entire original sequence. Terminated fragments are electrophoresed and the original sequence can be deduced. What improvement allowed for the automation of Sanger Sequencing? - ANSWERdye primer and dye terminator allowed for each nucleotide to be labeled by a different fluorescent dye What colors are each nucleotide labeled in Sanger sequencing? - ANSWERA- Green G-Yellow/Black C-Blue T-Red Does dye primer sequencing or dye terminator sequencing allow for the Sanger Sequencing reaction to occur in on reaction tube? - ANSWERdye terminating sequencing because the ddNTP's are labeled with dye vs. the primer which would