Isolate genomic DNA and break Into fragments. Deposit the beads Into a picotiter plate. Only one bead can fit Into each well. Fragment of genomic DNA Covalently attach oligonucleotide adaptors to the 5' and 3' ends of the DNA. Adaptors Denature the DNA Into single strands and attach to beads vla the red adaptors. Note: Only one DNA strand is attached to a bead. Add sequencing reagents: Primers, DNA polymerase, ATP sulfurylase, luciferase, apyrase, adenosine-5'- phosphosulfate, and luciferin. Sequentially flow solutions contalning À, T, G, or CInto the wells. In the example below, T has been added to the wells. PP, (pyrophosphate) Is released when Tis Incorporated Into the growing strand. Thymine nucleotides -DNA strand (shown In black) Is complementary to adaptor shown in red. * Adaptor Bead Emulsify the beads so there is only one bead per droplet. The droplets also contain PCR reagents that amplify the DNA. Primer (complementary to the adaptor shown in green) PP. + Adenosine-5'- phosphosulfate ATP sulfurylase ATP + luciferin Luciferase Light Light is detected by a camera In the sequencing machine. Unincorporated nucleotides and ATP are degraded Бy aругase. FIGURE 23.14 Pyrosequencing, an example of a newer highthroughput sequencing technology.
Isolate genomic DNA and break Into fragments. Deposit the beads Into a picotiter plate. Only one bead can fit Into each well. Fragment of genomic DNA Covalently attach oligonucleotide adaptors to the 5' and 3' ends of the DNA. Adaptors Denature the DNA Into single strands and attach to beads vla the red adaptors. Note: Only one DNA strand is attached to a bead. Add sequencing reagents: Primers, DNA polymerase, ATP sulfurylase, luciferase, apyrase, adenosine-5'- phosphosulfate, and luciferin. Sequentially flow solutions contalning À, T, G, or CInto the wells. In the example below, T has been added to the wells. PP, (pyrophosphate) Is released when Tis Incorporated Into the growing strand. Thymine nucleotides -DNA strand (shown In black) Is complementary to adaptor shown in red. * Adaptor Bead Emulsify the beads so there is only one bead per droplet. The droplets also contain PCR reagents that amplify the DNA. Primer (complementary to the adaptor shown in green) PP. + Adenosine-5'- phosphosulfate ATP sulfurylase ATP + luciferin Luciferase Light Light is detected by a camera In the sequencing machine. Unincorporated nucleotides and ATP are degraded Бy aругase. FIGURE 23.14 Pyrosequencing, an example of a newer highthroughput sequencing technology.
Human Anatomy & Physiology (11th Edition)
11th Edition
ISBN:9780134580999
Author:Elaine N. Marieb, Katja N. Hoehn
Publisher:Elaine N. Marieb, Katja N. Hoehn
Chapter1: The Human Body: An Orientation
Section: Chapter Questions
Problem 1RQ: The correct sequence of levels forming the structural hierarchy is A. (a) organ, organ system,...
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Is this a sequencing by synthesis method? Explain.
Transcribed Image Text:Isolate genomic DNA
and break Into fragments.
Deposit the beads Into a picotiter
plate. Only one bead can fit Into
each well.
Fragment of
genomic DNA
Covalently attach oligonucleotide
adaptors to the 5' and 3' ends of
the DNA.
Adaptors
Transcribed Image Text:Denature the DNA Into single
strands and attach to beads vla
the red adaptors. Note: Only one
DNA strand is attached to a bead.
Add sequencing reagents:
Primers, DNA polymerase,
ATP sulfurylase, luciferase,
apyrase, adenosine-5'-
phosphosulfate, and luciferin.
Sequentially flow solutions
contalning À, T, G, or CInto the
wells. In the example below, T
has been added to the wells.
PP, (pyrophosphate) Is released
when Tis Incorporated Into the
growing strand.
Thymine nucleotides
-DNA strand
(shown In black)
Is complementary
to adaptor
shown in red.
* Adaptor
Bead
Emulsify the beads so there is only
one bead per droplet. The droplets
also contain PCR reagents that
amplify the DNA.
Primer
(complementary
to the adaptor
shown in green)
PP. + Adenosine-5'-
phosphosulfate
ATP sulfurylase
ATP + luciferin
Luciferase
Light
Light is detected by a camera In the
sequencing machine. Unincorporated
nucleotides and ATP are degraded
Бy aругase.
FIGURE 23.14 Pyrosequencing, an example of a newer highthroughput sequencing technology.
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