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Flashcards in Exam 2 Deck (99):
1

What are bacterial terminator sequences?

-rHo dependent

rho factor used

-rHo independent

inverted repeat sequences transcribed form hairpin loops

 

2

Most bacterial transcripts are said to be collinear, what does this mean?

DNA directly encodes them. 

one-to-one correspondence between of bases between the gene and the mRNA transcribed from the gene

NO SPLICING!

2

What are homologs?

genes related to a second gene by descent from a common ancestral DNA sequence

3

What singles the beginning of DNA replication?

Helicase bindind to DNA and breaking the hydrogen bonds between the two strands to separate them. 

3

Who discovered transposable elements?

Barbara McClintock 

3

What are class 2 TEs?

-aka DNA transposons

-complete/autonomous TEs

-encode the protein transposase, required for insertion and excision

-never use RNA intermediaries

-

3

What interferes with L1 activity?

siRNAs interfere with L1 activity and are derived from the 5' UTR of L1 LTRs

3

What is the Medaka fish example of?

Example of how TEs affect gene regulatory regions and phenotypes

Tol2 transposon either hops out of fish cleanly (results in albinoism) or takes with it genetic material (pigment alterations). 

3

How were full length cDNA sequences selected for?

-Biotin capping the 5' end 

-Washing the cDNAs with an RNA digestign enzyme like RNase I

-Only cDNAs with biotin cap (full cDNAs) could be recovered

 

3

Beta-thalassemia is a result of ___

an insertion (point) mutation

3

What is eukaryotic termination?

-for RNA pol 1:

similar to Rho-dependent termination of bacteria

-for RNA pol 2:

complex, mRNA  might be synthesized well beyond coding region.

cleaved by a complex and then adenylated

for RNA pol 3:

similar to Rho-independent termination

4

What is the difference in bacterial/ eukaryotic promoters?

Bacteria- 3 elements

pribnow box (TATAA site)

consensus sequence TTGCCA

upstream element (AT-rich)

 

Eukaryotes- 7 elements

enhancer sequences (bind with activators to alter DNA structure and facilitate RNA pol 2 binding

need transcription factor

TATA box

 

5

In humans, what are the only active class of transposons (jumping genes)

The non-LTR class of retrotransposons (class 1 TEs)

 

5

What are nonautonomous TEs?

-TEs that require another TE to move

-They either lack the gene for transcriptase or retrotransposase, so must borrow from a nearby TE

-ex: McClintock's DS element

5

one form of leukemia is caused by ____

translocation (chromosomal) mutation

6

What characterizes class 2 TEs?

-inverted repeats on their ends to be recognized by transposase (ex: ACGCTA, TAGCGT)

-short direct repeats that flank inverted repeats and play a role in insertion. They are footprints left behind and sometimes alter gene expression. They are not actually part of TE, because they're left behind

7

What were the genetic crosses that McClintock performed?

What was the hypothesis of phenotype of the resulting progeny?

What actully occurred?

What was the conclusion?

C'C'bzbz- - x CCBzBzDsDs

To yield heterozygots:

C'CBzbzDs-

Expected:

all progeny would be colorless

Observed:

many kernels were colorless but some cells were purple or brown resulting in spots/streaks of color

Conclusion

A breakage at the Ds locus caused some cels to lose the C' and Bz that would yield colorless or brown phenotypes. Depending on when the breakage occured during development, there was more or less spotting

8

What are autonomous TEs?

-TEs that can move on their own

-ex: McClintock's AC elements

9

What are the minor/major grooves of the DNA double helix? What is their role?

Major/minor groove: caused by the asymmetric spacing of the antiparallel helix backbone

Their Role: role in binding of proteins that regulate gene transcription

10

Besides epigenetic silencing, how do TEs silence themselves?

siRNA, allows TEs to mediate their own silencing

siRNA: prevent transposition. double stranded RNA

11

How do siRNAs interefere with L1 activity?

5' UTR of L1 promoter encodes a sense promoter that transcribes L1 gene and an antisense promoter that transcribes an antisense RNA

The result is homologous sequence and hybridization and a double stranded RNA that can serve as a substrate for RNAi. 

12

some cancers are caused by ____

insertion (chromosomal) mutation

14

                                                                                                        What differentiates DNA from RNA?

DNA doesn't have a 2' OH group on the sugar molecule

14

Who is Kornberg?

-enzymologist and nobel prize winner

-described DNA replication as a tape recording of instructions for completing a task 

-discovered that DNA polymerase catalyzes DNA synthesis by mixing necessary ingredients for e-coli DNA synthesis in a test tube

14

How were okazaki fragments discovered?

-Pulse chase experiment

- DNA exposed to isotopic-nucleotides (pulse) and then regular nucleotides for varying periods of time (chase)

-short chases resulted in most radioactivity in "slow" DNA

-increasing length of chases resulted in radioactivity increase in "fast" DNA but not "slow" DNA

-shows that in short period of time okazaki fragments form but with more time they elongate as DNA replication continues

15

What are class 1 TEs? How do they differ from class 2 TEs?

-aka retrotransposons

-move through the action of RNA intermediaries

-do not encode RNA transcriptase, rather produce RNA transcripts and rely on reverse transcriptase enzymes to reverse transcribe RNA into DNA, which is inserted into target site

16

What are chemical mutations?

Free radicals (oxidizing agents) may modify nucleotides

Example: Dioxin intercalates between base pairs, disrupting DNA helix integrity 

Example: benzo[a]pyrene induces lesions at guanine sites in P53, lung cancer!

17

Why are RNA primers required at the origin of replication?

Because DNA can't bind de novo and can only add deoxyribonucleotides to the 3' OH group of an existing chain 

18

Human chromosomes range from _ o _ in length

50 to 250 million base pairs

19

What are transposons?

-Jumping genes

-DNA sequences that move from one location on the genome to another

19

What are the essential chemical ingredients in PCR?

- DNA polymerase

-deoxyribonucleotides

-essential salts (like Mg)

-DNA template

primers with exposed 3' OH groups 

20

What is the central dogma of molecular biology?

DNA->RNA-> proteins

First step transcription

Second step translation

21

Cri-du-chat syndrome is caused by ___

deletion (chromosomal) mutation

22

What is an example of a mutation "hot spot"?

regions of DNA with large numbers of trinucleotide repeats. 

This could lead to DNA polymerase slippage and alteration of the repeat sequence 

23

What is the length of a turn of DNA?

What is the width of the DNA double helix?

How many bases are in 1 turn?

length: 3.4 nm (34 A)

width: 2 nm (20 A)

1 turn: 10 nucleotide bases

24

What is the Ames test?

Test to determine if a chemical compound or any of the digested material derived from it is a mutagen 

24

What is a protein domain?

a region tha can adopt a 3D structure

25

What are two families of non-LTR retrotransposons?

-Line1 (L1) (LONG! LESS!)

-6 kb in length

-less copies in humans but longer so 15-17% of human genome

 

 

-Alu

-only a few hundred nucleotides

-more copies in humans

27

What is cDNA?

complimentary DNA synthesized from mRNA using reverse transcriptase

 

28

What is the role of DNA polymerase I?

Removes the RNA primer after DNA polymerase III has begnun and replaces it with nucleotides

29

What are two ways in which sequences are fixed

proofreading- correction during replication

mismatch repair-correction after replication

30

What is the role of DNA polymerase III?

does most of the elongation work 

adds nucleotides one by one to the 3' end of a growing single strand 

31

In what direction does DNA synthesis occur?

5' to 3' direction ONLY

32

What are the bonds that link the backbone of DNA and what are they made up of?

Phosphodiester bonds of the DNA backbone are made up of 5 carbon sugars and phosphate groups

33

What is a protein motif?

-short, conserved region of a protein

-10 to 20 contiguous residues

 

34

What are the sequences at the start and end of splice sites?

Start: GU

END: AG

35

What adds new nucleotides to an DNA strand during DNA replication?

DNA polymerase

36

What are the two types of class 1 TEs?

Class 1 TEs: retotransposons

Type 1: LTR retrotransposons

-Have long terminal repeats on both ends

Type 2: non-LTR retrotransposons

-do not have long terminal repeats at both ends

36

What are orthologs?

genes in different species that evolved from a common ancestral gene

37

What results in sickle-cell anemia?

A point mutation in a nucleotide converts the sixth amino acid from glutamic acid into valine 

38

How is genetic information encoded in DNA?

genetic information is encoded in the sequence of bases attached to the 1' carbon of the sugar molecule of the DNA backbone

39

What are connected to the carbons of the sugar molecule in DNA?

1' carbon: OH group is replaced by nucleotide bases

2': in RNA, has an OH group. In DNA, it does not

3': a phosphate group attaches here

4': attached to the 5' carbon

5' carbon: 5' phosphate group 

40

What is an example of environmental mutation?

UV mutations induce hydrolysis of cytosine causes it to mispair to adenine

(creates a C-T fingerprint type mutation)

UV also induces covalent bonds between adjacent pyrimidines

42

What were some other discoverys by McClintock?`

-Additional discoveries showed an element Ac that was necessary for phenotypic effects of Ds

-McClintock had trouble mapping Ac and Ds, noting that they changed locations

 

44

T/F: Scientists have identified a polymerase that can add nucleotides to the 5' end

FALSE

45

Why is maize ideal for genetic analysis?

Each kernel is a unique embryo so many different phenotypes can be scored on a single ear of corn 

46

How were TEs discovered?

-Barabara McClintock

-Studying maize

-studying phenotypic system of maize (triploid endosperm  that is the colored protein coating and diploid zygote

-Four maize genes studied: C' (dom, colored inhibitor)/ C (rec, color shown), Bz (dom, color purple)/ bz (rec, color brown), Ds (location on Ch9 where breakage occurs), As (unknown factor that impacts Ds expression)

Ds observed due to unexpected phenotypes in maize genetic crosses

48

What is the difference between purines and pyrimidines?

Purines: composed of two rings (guanine and adenine)

pyrimidines: composed of one ring (thymine and cytosine)

49

What is a single-nucleotide polymorphism?

A single base pair alteration that is common in populations

Basically, any location where at least two sequences are found and are prevalent in at least 1% of population

50

How are complimentary bases connected?

Complimentary nucleotides are hydrogen bonded to each other. 

51

Where does translation occur?

When does it begin?

In ribosomes

Once initiation factor proteins bind to small subunit of ribosome

Methionine carrying tRNA binds to mRNA near AUG start

Methionine is the first amino acid encoded in any new protein 

methionine not removed in MK case

53

cystic fibrosis is a result of ___

deletion (point) mutation

54

Optiz Kaveggia syndrome is a result of _____

An insertion (chromosomal mutation)

55

What is the role of DNA ligase

seals the bond between two adjacent nucleotides. 

This "nick" is due to removal of RNA primer and left behind by DNA polymerase I and RNase H. 

56

Which complimentary nucleotides form 2 hydrogen bonds and which complimentary nucleotides form 3?

2 hydrogen bonds: AT

3 hydrogen bonds: GC (hence why they form a stronger bond)

57

What is topoisomerase?

reduces torsional strain caused by DNA unwinding

58

What are two types of sponatenous mutations?

1. deamination

loss of an amino group in a nucleotide

typically if cytosine, repair mechanism fixes this unless cytosine is methylated. Then it is read as a thymine

(CpG islands frequent targets for methylation and mutation)

2. depurination

purine is removed so polymerase can't determine what nucleotide to add to new strand. 

generally polymerase adds an adenine (sometimes guanine)

60

What is the result of an expanding trinucleotide repeat?

The normal number of repeated trinucleotides is expanded

results in fragile X, huntington's disease

61

What catalyzes splice reactions and what type of reactions are involved?

snRNPs 

transesterification bonsd guanine and adenine

3' and 5' end of intron connected via transesterification as well 

62

What is a transversion mutation?

A->T

T->A

C->G

G->C

63

What is DNA cloning?

-DNA clonin involves the cloning of expressed DNA genes via a reversal of the central dogma of molecular biology

 

-An mRNA transcript is transcribed into DNA via reverse transcriptase 

64

What is the difference between a non-sense and a mis-sense mutation?

non-sense: changes AA sequence to premature stop codon

mis-sense: changes AA sequence

66

What are the differences between eukaryotic and prokaryotic replication?

eukaryotic:

-many replication origins

-DNA organized into nucleosomes with histones

-different set of polymerases (polymerasa beta and polymerase gamma instead of polymerase III)

-13 polymerases discovered to date

-chromosomes are linear

-gap in newly synthesized DNA ends protected by telomeres

 

prokaryotic:

-single replication origin (oriC)

-chromosomes are circular

67

where does the splicing process occur?

in spliceosomes in nucleus ?

68

What experimental evidence proved the existence of reverse transcriptase?

1. The DNA polymerase only incorporated deoxyribonucleotides, not ribonucleotides, into its product.

2. The product itself "behaved" like DNA--in other words, it was sensitive to treatment by deoxyribonucleases but not ribonucleases.

3. The RNA itself was the template, as shown by the fact that treatment of virions with ribonucleases destroyed the ability of the polymerase to incorporate radioactively labeled nucleotides.

70

How do TEs drive the evolution of genomes?

-help repair double-strand breaks

-exon shuffling

-translocation of gene sequences

 

71

What is gene amplication?

An example of copy number variants 

It's a result of the number of tandem copies of a locus being increased

in some breast cancers 

72

Transposable elements make up how much of human and maize genome

-50% of human genome

-90% of maize genome

74

What is meme?

searching for common motifs in unaligned sequence

75

What epigenetic defense mechanisms can silence TEs?

chromatin remodeling

chromatin becomes so constricted in some areas that transcription factors can't bind

DNA methylation (heterochromatin)

In Maize for example: wild-type sequences are methylated. In mutated maize, transposons are transcribed. 

miRNA

Used in gene regulation: cause histone modification and DNA methylation, bind to mRNA

 

77

What is exon shuffling? Why does it occur?

Exon shuffling occurs becaue TEs do not always excise perfectly from sequence. The result is that two previously unrelated exons are are juxtaposed, potentially creating new gene products

 

 

78

79

What is oriC?

The replication origin

a base-pair sequence of nucleotides

80

What is a transition mutation?

Mutation of purine to purine or pyrmidine to pyrmidine

So:

T->C (pyrimidine)

A-> G (purine)

81

What are the 3 types of RNA  polymerases?

In eukaryotes:

RNA Pol 1: ribosomal RNAs

RNA Pol 2: responsible for mRNA synthesis from template DNA

RNA Pol 3: responsible for small RNA, tRNA synthesis

82

Role of RNA primase?

inserts starter of RNA nucleotides at trancription start site

also attaches RNA primers to lagging strand to yield okazaki fragments

83

What is the role of RNase H?

Removes the RNA primer after DNA polymerase III has begnun and replaces it with nucleotides

84

What is the usefulness of cDNA libraries?

can be used to identify genes that are expressed differently in different types of tissues or different stages of development

cDNA libraries give a snapshot of gene activity since only  genes that are expressed and transcribed into mRNA can be cloned

86

What is prosite?

a database of protein patterns that can be searched by either regular expression patterns orsequence profiles

88

What is phi blast

searching a specific protein sequence pattern with local alignments surrounding the match 

89

What is the difference between the 5' and 3' end of the DNA double helix?

5' end: has a free 3' phosphate group (negatively charged)

3' end: has a hydroxyl group on the 3' carbon of the sugar

91

How does replication of the lagging strand occur?

RNA primase creates an RNA primer with 3' OH so that DNA replication can begin. 

DNA polymerase III adds deoxynucleotides in the 5' to 3' direction following the 3' end of the primer

DNA polymerase I replaces DNA polymerase III and replaces RNA with DNA after exonuclease removes RNA primer

DNA ligase binds 3' OH of growing strand with 5' phosphate of the existing strand by eliciting formation of a phosphodiester bond. 

93

What are paralogs?

genes related by diplication within a genome 

94

What is the difference between the leading and lagging strand?

DNA polymerase can only add new nucleotides to a free 3' end of a growing chain. 

Leading strand: replication occurs continuously in the 5' to 3' direction

lagging strand: to provide a 3' OH, RNA primase attaches to the DNA and synthesizes a short RNA primer. DNA polymerase III then adds deoxynucleotides to the 3' end of the primer. DNA polymerase I replaces the RNA with DNA. The enzyme DNA ligase forms a phosphodiester bond betwen the 3' OH of the growing strand and the 5' phosphate in front of it. 

95

What is the role of SSBPs?

singal stranded binding proteins keep separated DNA strands from reattaching 

96

What do jumping genes do?

Examples?

-May result in mutation when they insert into a gene

-Example: L1 into Factor VIII results in hemophilia

-Example 2: L1 in APC genes in colon cancer

-L1 transposes in mammalian somatic cells, which could play a role in disease development. 

97

Where does helicase split the DNA molecule apart?

At the start of the replication fork, typically areas rich in A and T because they're only connected by two hydrogen bonds 

98

What are the 4 steps to replication?

1. initiation

2. unwinding

3. primer synthesis

4. elongation

 

99

What is a silenced TE?

Why are some TEs silenced?

-Silenced TE: does not produce a phenotypic effect (opposed to L1, most TEs appear to be like this)

-Inactive due to:

  • mutations that affect ability to move around chromosomes
  • kept inactive by epigenetic defense mechanisms