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BMME 485 > Exam 1- Lecture 6 > Flashcards

Exam 1- Lecture 6 Flashcards

1
Q

What are linkers/adapters

A

short pieces of DNA with a known sequence

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2
Q

Why are linkers/adapters used

A

Because the end of DNA fragments are unknown after they have been sheared through sonication

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3
Q

What is a barcode or index sequence

A

sequence unique to the sample of DNA adaptor

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4
Q

What is multiplexing

A

allows for multiple samples of DNA to be analyzed at the same time

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5
Q

What is read depth

A

the number of sequences that map to a region in the genome

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6
Q

What is read depth used for

A

to ensure that a change is not an error in the sequencing process

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7
Q

In 454 sequencing where are beads attached to

A

beads via adaptors

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8
Q

What is present in the beads in 454 sequencing

A

small DNA oligonucleotides complementary to the adaptor

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9
Q

Once the fragments with adaptors are placed into beads what happens

A

one DNA fragment will attach to single bead

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10
Q

In Illumina sequencing, where are DNA fragments added

A

to the surface of a flow cell

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11
Q

What is on the surface of a flow cell for Illumin sequencing

A

DNA primers complementary to the adaptor on each of the DNA strands

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12
Q

For 454 sequencing, what is used to create multiple copies of the single piece of DNA

A

emulsion PCR

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13
Q

How does emulsion PCR work

A

emulsion of oil and water
one bead into one water droplet
water contains deoxynucleotides, primers complementary to adaptors, and Taq DNA polymerase
DNA amplified normally inside water
bead coated with identical copies of DNA

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14
Q

What does Illumina sequencing use to create multiple copies of DNA

A

bridge amplification

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15
Q

How does bridge amplification work

A

flow cells are incubated with deoxynucleotides and DNA polymerase
primers are attached to flow cell
DNA anneals to primer on surface–>forming bridge
amplified and release to form cluster of fragments

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16
Q

One cluster of identical strands are produced in Illumina or DNA sequencing, what happens

A

strands are denatured into single stranded DNAs for sequencing

17
Q

How does 454 sequencing work

A

beads separated into picotiter plate
primer annealed to adaptor sequence
one of four deoxynucleotides and DNA polymerase floated over wells
if well has complementary base pair, nucleotide added to primer, pyrophosphate released
flash of light occurs and is recorded

18
Q

How does Illumina sequencing work

A

After primer is annealed to adaptor sequence
all four nucleotides that are fluorescent are added to flow cell at same time
wavelength of light released at each flow cell is recorded
fluorescent dye terminator is removed and another batch is added to

19
Q

What is a read

A

single piece of sequence information

20
Q

What is contained in the well in 454 sequencing

A

luciferase and sulfurylase

21
Q

How does Ion Torrent sequencing work

A

microwell filled with template DNA strand
filled with deoxyribonucleotide triphosphate
if dNTP is complementary to leading template nucleotide
dNTP is added to growing complementary strand
hydrogen ion is released
triggers ion sensor

22
Q

In oxford nanopore sequencing, what is different about the flow cells

A

the flow cells contain nanopores embedded with electro-resistant membrane

23
Q

How does oxford nanopores read DNA sequences

A

as DNA passes through nanopores, electrical current changes as different nucleotides pass through, resulting signal is decoded to provide specific DNA sequences

24
Q

What does PacBio use as a signal for sequencing by synthesis

A

Light

25
Q

What is multiplexing

A

barcode index sequence gives ability to pool samples and run them together at same time

26
Q

Ion torrent is

A

sequencing by synthesis

27
Q

PacBio is

A

sequencing by synthesis

28
Q

Oxford nanopore is not

A

sequencing by synthesis

BMME 485 (15 decks)

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