trans Flashcards
(124 cards)
1
Q
_____ – in contrast to bacterial cells, all eukaryotic cells
contain membrane-bound organelles.
A
Organelles
2
Q
Cell membrane functions as shell of each cell in the body; contains a hydrophilic and hydrophobic
surface; encloses contents of the cell.
A
lipid bilayer
3
Q
Membrane bound organelles within the cell are encased in a ____
A
lipid bilayer
4
Q
Structure containing the majority of genetic
material within the cell.
A
nucleus
5
Q
_____ – genetic material of the cell; organized into
chromosomes.
A
DNA
6
Q
Ø Numerous within each cell; number varies by cell type.
Ø Energy generation occurs within _____
Ø Oxidative phosphorylation
Ø Small amount of DNA
A
mitochondria
7
Q
Ø Structure continuous with the nucleus; contains no DNA &
Ø Site of protein synthesis
A
Endoplasmic reticulum
8
Q
Ø Protein modification &
Ø Protein transport to the cytosol
A
Golgi apparatus
9
Q
Ø Protein modification &
Ø Protein transport to the cytosol
A
Golgi apparatus
10
Q
Ø Function in removal of waste products within the cell
A
Lysosomes and peroxisomes
11
Q
Structure of DNA
a. Purine and pyrimidine bases: A, T, G, C
b. A ______ backbone links the bases together to form a chain.
c. A ______r is also contained at each base position.
d. Double helix. Two DNA chains are _______bonded to one another between A and T or C and G. The two joined, antiparallel strands then twist into a double helix.
e. ______ Along the DNA strands are specific, short regions that encode for ___.
A
phosphodiester
deoxyribose suga
hydrogen
Genes.
12
Q
Chromosomes – human DNA is further arranged into ___
pairs of matched chromosomes
A
23
13
Q
– the DNA strands are wound around histone proteins and these coil together to form a tightly packed _____
A
Chromatids
14
Q
two sister chromatids are joined at
the _____. These two joined chromatids form a chromosome.
A
centromere
15
Q
the ends of chromosomes contain
repetitive sequences of a specific 6-base sequence: ______. This repeated sequence can be many
hundreds of base pairs long.
_______protect
genes at the ends of chromosomes from normal damage and shortening during DNA replication. They are also thought to be involved in the aging process.
A
TTAGGG
Telomeres
16
Q
Ø Extrachromosomal, circular pieces of DNA first identified in bacterial cells
Ø Confer antibiotic resistance to bacteria
Ø Engineered in the laboratory to clone specific pieces of human DNA
A
Plasmids
17
Q
Ø Semiconservative process
Ø Strands are separated by enzymes and new nucleotides added to form a new strand using the original strand as a _____. Replication is carried
out by _______.
Ø Result is the generation of two identical strands of
DNA, each of which contain one strand from the original duplex.
Ø Carried out in the ____ direction. One stand is
directly copied from its template. Other strand is
replicated in small pieces known as Okazaki
fragments. These fragments are later joined by DNA
ligase to form a contiguous strand of DNA.
A
DNA Replication
template
DNA polymerases
3’ to 5’ direction
18
Q
RNA
- Classes
a. ______ – defines amino acid to be incorporated into a polypeptide
b. _______ – forms a portion of the ribosome; participates in protein synthesis
c. ______ – template from which a polypeptide or protein is made
A
tRNA
rRNA
mRNA
19
Q
_______ – process of converting genetic code contained within the DNA into an RN copy. After this process, the mRNA copy undergoes specific ______ modifications to prevent degradation, function in controlling the rate of protein synthesis, repair misincorporated nucleotides, or increase genetic diversity.
A
Transcription
post-translational
20
Q
_______ – addition of long sequence of A nucleotides to the 3’ end of mRNA
A
Polyadenylation
21
Q
addition of a methyl group to the 5’ end of mRNA
A
Capping
22
Q
______ – inserting, deleting, or editing nucleotides after synthesis
A
splicing
23
Q
Ø Represented by a triplet codon sequence
Ø Basic components of proteins
Ø Contain an amino and carboxyl group
Ø Twenty _____ found in proteins
A
Amino acids
24
Q
Ø Chain of amino acids
Ø Alpha-helix or beta-sheet
Ø Component or subunit of a mature protein
A
Polypeptides
25
Ø All are proteins
Ø Catalyze specific chemical reactions within or outside of the cell
Ø ______ proteins are common examples of enzymes
Ø Degradation of cellular components
Ø Cell division
Ø Also produced commercially for use in the laboratory
Enzymes
| DNA synthesis proteins
26
________– most common type of molecular diagnostic specimen. Blood is most commonly collected in ______ tubes. ______ is a potent
inhibitor of PCR and is not used in molecular diagnostics. Blood should be stored at ____ until processing.
Whole blood
purple-top
Heparin
4*C
27
______ – specimens should be frozen immediately upon receipt or held in cell culture medium at _____
until processing
Tissue
| 37*C
28
______ – often used for remote site testing; collection of DNA specimens for parentage testing.
Swabs should be ____ before shipment to the laboratory.
Buccal swabs
| air-dried
29
_____ – urine, feces, sputum, CSF, genital swabs, semen, sections of formalin-fixed paraffinembedded tissues, bone, hair, fingernails usually
confined to ______specimens
Other specimens
| forensic
30
DNA isolation
_______ – provide ease of use and high degree of reproducibility. Yields _____amounts of DNA. DNA is usually very pure. Most often used to isolate DNA from blood. This is the most commonly encountered clinical DNA isolation method.
Manufacturer (Qiagen) kits
smaller
31
DNA Isolation
_______– fastest method. Low quality, low mass DNA recovery. Usually used to isolate DNA from buccal/FTA cards.
Quick-extract solutions
32
RNA isolation
a. _______ – gold standard. Yields a large mass of high quality RNA. Can be used
on any specimen type.
b. ________ – provide ease of use and high degree of reproducibility. Preferred method for clinical RNA isolation.
c. ________. Increase recovery of mRNA in specimens.
Acid phenol/LiCl salt isolations
Manufacturer kits
Poly-A enrichment kits
33
Spectrophotometry
Ø ___ value is used to calculate nucleic acid mass.
Ø DNA A260/280 ratio between _______ – indicative of DNA quality sufficient for clinical manipulation. Lower ratios indicate impure specimens.
Ø RNA A260/280 ratio between _____– indicative of RNA quality sufficient for clinical manipulation. Lower ratios indicate ____ specimens.
A260
1.8 and 2.0
1.9 and 2.1
impure
34
1. Target amplification
a. _______ – method by which almost all molecular diagnostic assays are performed. It is a three-step process consisting of melting the duplex DNA strands, annealing the primers, and extending the new fragment to yield a specific, short DNA product. This process is repeated 25 to 40 times to theoretically yield trillions of copies
from one starting copy of DNA.
Ø _____ – short segments of synthetically manufactured DNA used to define a specific
position to be copied in the PCR process. Each PCR reaction requires two primers – one on either side of the fragment to be amplified.
Ø _______ – salt required as a cofactor for the polymerase enzyme used to carry out PCR.
Polymerase chain reaction (PCR)
Primers
Magnesium chloride
35
_______– enzyme used to generate new DNA copies in the PCR process. A thermostable enzyme, meaning that it will not degrade at very high temperatures used in the PCR process.
Ø_____ – containing potassium chloride, sodium chloride, and other additives are also included in the reaction to stabilize the enzymes and DNA during the PCR process.
Taq polymerase
| Buffers
36
_______ – individual building blocks of the DNA strand.
dNTPs (A, T, G, C)
37
_______ – a very small mass of this specimen from the patient to be tested is also added to the reaction. The PCR process only requires ______ quantities of input DNA.
Genomic DNA
| femtomolar
38
________ – an alternative method to PCR that is not based upon direct amplification of the genomic DNA template.
Transcription-based amplification (TMA)
39
_______ -- another method that does not rely on direct amplification of the template DNA strand. Two commonly encountered clinical examples of this strategy
include:
a. ___________
b. _________ – used clinically to detect deletions of multiple entire exons in very long stretches of DNA representing large genes.
c. _____________
Probe amplification
Ligase chain reaction
Multiplex ligation-dependent probe amplification (MLPA)
Strand displacement amplification
40
_______ – another method of detecting a DNA target without direct amplification of the template DNA. It relies on detection of substrates bound to the target nucleic acid.
a. Branched DNA amplification
b. Hybrid capture
Signal amplification
41
Method used in many applications within the molecular diagnostics laboratory. Used to fractionate DNA at sequence-specific locations using restriction enzymes. ______ is used to generate size standards and to prepare genomic DNA for analysis by Southern
blotting and hybridization.
Restriction digestion
42
______– method to analyze DNA contained in an entire genome through a process of restriction digestion, electrophoresis, transfer to a membrane, and hybridization to a sequence-specific probe.
Southern blot
43
_______– analogous to the process of Southern blotting, but is performed on RNA specimens.
Northern blot
44
_______ – small fragments of DNA used to detect a specific gene or mutation in the process of Southern or Northern blotting
Probes
45
_______– conditions, i.e. salt concentration, temperature, time and buffer composition, that control the stringency of the hybridization procedure.
Hybridization dynamics
46
A method used to separate DNA fragments onto a solid matrix based upon size
Gel electrophoresis
47
1. ___________ – most often performed to resolve DNA fragments larger than 100 bp in size.
2. __________ – method of choice to resolve small DNA fragments. Very high concentrations of polyacrylamide matrix can resolve down to 1 bp differences in size.
3. __________ – modification of polyacrylamide gels used in automated DNA sequencing and genotyping platforms.
Agarose gel electrophoresis
Polyacrylamide gel electrophoresis
Capillary gel electrophoresis
48
Quantitative Real-time Polymerase Chain Reaction
A PCR method starting the mRNA as a ____. This method is capable of quantifying the copy number of mRNA transcripts present in a specimen.
1. ________ – enzyme used to make a copy DNA, or cDNA, from the mRNA transcript contained in a specimen. This process is referred to as cDNA synthesis.
2. ________– general term for the PCR method used to observe the production of new DNA fragments in real time on an automated platform.
a. _____ – dye often used in quantitative real-time PCR assays to detect newly generated DNA fragments
b. ______ – alternate system of
amplification and detection of fragments in quantitative real-time PCR using dye-labeled probes.
```
template
Reverse transcriptase
Real-time PCR
SYBR green
TaqMan chemistry
```
49
______ - Allows determination of each base pair in a single strand of DNA.
__________ – original method of DNA sequencing; basis of all DNA sequencing used in the clinical environment today. It is based upon the incorporation of dideoxy nucleotides in random positions in the newly synthesized DNA strand.
_________– method by which DNA
sequence is determined based upon incorporation of a dye-labeled dideoxy nucleotide and resolution on a capillary electrophoresis instrument. Each dideoxy
nucleotide is labeled with a specific dye and when incorporated, is excited by a laser and detected by a CCD
camera on an automated platform.
DNA Sequencing
Sanger dideoxy sequencing
Fluorescent DNA sequencing
50
Technologies are the newest area of use in molecular diagnostics. They fall in two general categories:
1. ___________ – designed to
detect single base or other small changes in DNA sequence across the entire genome simultaneously. Many thousands of positions can be interrogated in one experiment.
2. _________– designed to asses global changes in expression of specific genes in a given tissue at a given time or under a specific set of treatment parameters.
Single nucleotide polymorphism arrays
| Gene expression arrays
51
```
_________
Ø Trinucleotide repeat expansion
Ø X-linked
Ø Mental retardation
Ø Primary affects males
```
Fragile-X
52
Ø Deletion of entire exons
Ø Dystrophin gene
Ø X-linked
Ø Muscle degeneration
Duchenne muscular dystrophy
53
Ø Trinucleotide repeat expansion with anticipation; the worsening of clinical symptoms with increasing repeat size in successive generations
Ø Autosomal dominant
Ø Males and females
Ø Multisystem disease
Myotonic dystrophy
54
Ø Many individual mutations in the _____may give rise to CF alone or in combination
Ø ______ mutation accounts for over 70% of CF cases
Ø Autosomal recessive
Although it primarily affects the lungs, it is a multisystem disease
Ø Most common genetic disease in Caucasians
CFTR gene
| Delta F508
55
DNA isolation
_________ – gold standard. Yields a _______of high quality DNA. Can be used to isolate DNA from any specimen type. This
method normally requires a large mass of starting material.
Phenol:Chloroform:Isoamyl alcohol method
large mass
56
_______________ – commonly performed assay in molecular diagnostics. Determination of the effectiveness of a therapy or the tracking of a patient’s disease status based upon the presence of
specific mRNA transcripts known to cause leukemia or
lymphoma.
a. ___________ – presence of
Philadelphia chromosome or 9;22 translocation. The
transcript can be detected by quantitative real-time
PCR and the size determined by fluorescent capillary
electrophoresis.
Minimal Residual Disease Detection
| Chronic myeloid leukemia (CML )
57
Chronic myeloid leukemia (CML )– presence of
Philadelphia chromosome or 9;22 translocation. The
transcript can be detected by ________ and the size determined by ___________
quantitative real-time PCR
| fluorescent capillary electrophoresis.
58
_______ – presence of a specific T or B cell clone
originating from a lymphoma. The clone is detected using
_____and _________
Clonality
PCR
fluorescent capillary electrophoresis.
59
__________– a genetic phenomenon
observed in cells undergoing changes in specific cells. The
expansion of short repetitive stretches of DNA to much
longer stretches that change gene expression within the
cell.
Microsatellite instability
60
______– hereditary form of colon cancer that is
| marked by a microsatellite instability
HNPCC
61
__________– nature of some tumors to exhibit
loss of function of one allele in a specific gene in a cell in
which the other gene was already inactivated. The most
common example of a cancer caused by this process is
______, in which expression of a tumor
suppressor gene is lost
Loss of heterozygosity
retinoblastoma
62
_______________ – can be determined
| through genotyping of DNA or gene expression analysis.
Drug metabolism and sensitivity
63
____________– performed to determine drug sensitivity. Test is carried out on a microarray platform.
Cytochrome P450 genotyping
64
Bacterial identification. The following organisms can be
easily detected,___________identification, using ____:
a. Chlamydia trachomatis
b. Neisseria gonorrheae
c. MRSA
d. VRE
qualitative
| PCR
65
```
Viral identification. Performed in ________ manner.
Following may be ____ detected:
a. Viral load – _____
i. HCV
ii. HIV
iii. CV
```
quantitative
66
There are a number of ways by which human identification
may be performed. The most widely used system is based
upon determination of the length of repetitive DNA in specific
regions of chromosomes across the human genome. This is
termed ______________
DNA-based human identification.
67
___________ – not as diverse as those observed
using other systems of identification. This is an older
system of human identification.
HLA Polymorphisms
68
__________ – first DNA-based identification system.
Based upon polymorphism in the length of restriction
enzyme-cut products across the genome. It utilizes
Southern blotting and hybridization as a method of
laboratory testing.
RFLP Analysis
69
It is most widely used system for identification of
individuals
Ø Very high power of discrimination
Ø ________are typed and compared to the
CODIS database for criminal investigations
Ø _________ typing of older specimens or cold cases
utilizes this system when the specimens are in
acceptable condition
Ø ________uses the same STR system as those
used in the CODIS allele database
Ø _____ engraftment is a clinical application of
this chemistry. STR analysis may be performed to
track donor versus recipient DNA components
following transplant.
Ø Other uses of DNA-based human identification
include QC of _______ specimens to ensure that a
patient matches a specific specimen.
STR Analysis
```
Common alleles
Forensic DNA
Parentage testing
Bone marrow
histological
```
70
____________ – identification of individual
based upon their mitochondrial haplotype.
Ø Primarily used on degraded specimens
Ø Only capable of determining maternally inherited
alleles with this system, because mitochondria are
passed from mothers to all offspring
Ø Often used in cold cases, because there are many
hundreds of copies of mitochondrial DNA present for
every one copy of nuclear DNA. This allows for a
greater possibility of success in analysis.
Mitochondrial DNA analysis
71
```
___________– newer system for human
identification
Ø Only capable of determining paternally inherited
alleles
Ø System only used to type males
Ø Based upon repeat length polymorphism
```
Y-chromosome analysis
72
__________– coined by James Watson; referred to the
biology of DNA; study of nucleic acids. Handling and analysis of
nucleic acids, DNA and RNA
Molecular Biology
73
DNA
Macromolecule of C, N, O, P, and H atoms; assembled in
units of nucleotides composed of a__________and a __________
Nitrogen bases: adenine, cytosine, guanine, thymine.
They are attached to a deoxyribose sugar which forms a
polymer with the deoxyribose sugars of other
nucleotides through a ______ bond.
phosphorylated ribose sugar
nitrogen base.
phosphodiester
74
NUCLEOTIDES
• Each nucleotide consists of a _________ The first
carbon of which is covalently joined to a _____ base
and the fifth carbon to a _____ moiety.
• Can be converted to nucleosides by _______.
• ________: a nitrogen base bound to an
unphosphorylated sugar.
• If the ribose sugar is _______ the molecule is a
nucleoside mono-, di-, or triphosphate or a nucleotide.
• The hydroxyl group on the ____ carbon is important for
forming the _________ that is the backbone
of the DNA strand.
• _______ – double ring structure; _______.
• ________ – single ring structure; ______.
• ________________________are the key to the
specificity of most nucleic acid-based test used in the
molecular laboratory.
• _________________ is also how the
information held in the linear order of the nucleotides is
maintained. As DNA is polymerized, each nucleotide to
be added to the new DNA strand, hydrogen bonds with
the complementary nucleotide on the parental strand
(A:T, G:C)
```
five-carbon sugar; nitrogen; phosphate
hydrolysis
Nucleoside
phosphorylated,
third; phosphodiester bond
Purines; guanine, adenine
Pyrimidines; thymine, cytosine
Hydrogen bonds between nucleotides
Specific hydrogen bond formation
```
75
NUCLEIC ACID
• A macromolecule made of nucleotides bound together
by the _______ and __________ on their sugars.
• A nucleic acid chain grows by the attachment of the _________ group of an incoming nucleotide to the ________ of the last nucleotide on the growing chain.
• Addition of nucleotides in this way gives the DNA chain
a polarity; that is, it has a 5’ phosphate end and a 3’
hydroxyl end. We refer to DNA as oriented in a _________, and the linear sequence of the nucleotides, by
convention, is read in that order.
• The bases are positioned such that the __________
chain that connects them (sugar-phosphate backbone)
is oriented in a spiral or helix around the nitrogen bases.
• The DNA double helix represents two versions of the
information stored in the form of the order or sequence
of the nucleotides on each chain. The sequences of two
strands that form the double helix are ________,
not identical.
• They are in ________orientation, with the 5’ end of
one strand at the 3’ end of the order. Formation of
hydrogen bonds between two complementary strands
of DNA is called ________. Single strands of DNA with
identical sequences will ___ hybridize with each other.
```
phosphate; hydroxyl groups
5’ phosphate; 3’ hydroxyl group
5’ to 3’ direction
sugar-phosphate
complementary
antiparallel
hybridization
not
```
76
DNA REPLICATION
• As DNA synthesis proceeds in the 5’ to 3’ direction, __________, the enzyme responsible for polymerizing the nucleotide chains, uses a guide or template, to determine which nucleotides to add to the chain.
• _____________ is the key to maintaining the sequence of the nucleotides in DNA through new generations. It is important that this information, in the form of the DNA sequence, be transferred faithfully at cell division.
• DNA is first ______ from the helical duplex so that
each single strand may serve as a template for the
addition of nucleotides to the new strand.
• The new strand is then _______ by hydrogen bonding
of the complementary incoming nucleotide to the
nitrogen base on the template strand and then a
nucleophilic attack of the deoxyribose 3’ hydroxyl
oxygen on a phosphorus atom of the phosphate group
on the hydrogen-bonded nucleotide triphosphate.
• _________ is released with the formation of a
phosphodiester bond between the new nucleotide and
the last nucleotide of the growing chain. The duplicated
helix will ultimately consist of one template strand and
one newly synthesized strand.
• The antiparallel nature of duplex DNA and the requirement for the DNA synthesis apparatus to read the template strand in a 3’ to 5’ direction _____ consistent with copying of both strands simultaneously in the same direction.
```
DNA polymerase
Semi-conservative replication
unwound
elongated
Orthophosphate
are not
```
77
________– small fragments are the key to explaining how both strands of the parent helix are not copied in the same way.
Okazaki fragments
78
__________ – DNA replication proceeds in a
| continuous manner on the 3’ to 5’ strand
Leading strand
79
__________– 5’ to 3’ strand is copied in a
discontinuous manner from the leading strand’s
replication apparatus which jumps ahead a short
distance on the 5’ to 3’ strand and then copies backward
toward the replication fork.
Lagging strand
80
DNA cannot be synthesized _____; a preceding base
| must be present to provide they hydroxyl group.
de novo
81
______ – RNA synthesizing enzyme that catalyzes the
synthesis of short RNA primers required for priming DNA
synthesis. Must work repeatedly on the lagging strand to
prime synthesis of each Okazaki fragment
Primase
82
_____ – unwinds and untangles the double helix at the
| replication fork
Helicase
83
________ – relaxes the supercoiled DNA
Topoisomerase
84
_______– synthesizes the new DNA strand; also
| proofreads and corrects some errors
Polymerase
85
_______ – rejoins two DNA strands into a double helix and
| joins Okazaki fragments of the lagging strand
Ligase
86
_________ – used to manipulate DNA in vitro, for instance,
to make stepwise deletions in linearized DNA or to
modify DNA ends after cutting with restriction enzymes
Nuclease
87
_______ – catalyzes the addition of methyl
groups to nitrogen bases, usually _____ and ________
in DNA strands
Methyltransferase
| adenines; cytosines
88
Mispair base removed by _______. DNA polymerase tries a second time
exonuclease
89
It is similar to DNA, differs in sugar moieties and in one
nitrogen base component. _____ and _____,
respectively.
• Almost all ______ do not have complementary
partner strands, they are not completely single
stranded. Through __________, RNA species
fold and loop upon themselves to take on a doublestranded character that is important for their function.
RNA can also pair with complementary single strands of
DNA or another RNA and form a double helix.
RNA
Ribose; uracil
RNA strands
internal homologies,
90
• The copying of one strand of DNA into RNA by a process
similar to that of DNA replication.
• Activity is catalyzed by _______, which occurs
mostly in _____.
Transcription
RNA polymerase
interphase
91
TRANSCRIPTION
_________– synthesis of mRNA, the type of RNA that
carries genetic information to be translate into protein.
Polymerase II
92
Transcription _______
Ø Regulated in all organisms so that genes are
transcribed as required by specific cell types.
Ø ________ – where RNA polymerase and its
supporting accessory proteins assemble on DNA
Ø _________ of transcription or RNA synthesis
greatly outnumber DNA initiation sites in both
prokaryotes and eukaryotes.
Ø ______ in prokaryotes and eukaryotes
differ and work with different supporting proteins
to find and bind to DNA in preparation for
transcription.
Initiation
Promoter
Initiation sites
RNA polymerases
93
Transcription ______
Ø RNA polymerases in both eukaryotes and prokaryotes synthesize RNA using the base sequence of one strand of the double helix as a guide.
Ø The DNA double helix is locally _____ into single strands to allow the assembly and passage of the transcription machinery, forming a _______.
Ø RNA synthesis does not require _____. Subsequent ribonucleoside triphosphates retain only the _______, the one closest to the ribose sugar.
Elongation
unwound
transcription bubble
priming
alpha phosphate,
94
Transcription __________
Ø Accomplished in some genes by interactions
between RNA polymerase and nucleotide signals
in the DNA template.
Ø _____ – helicase enzyme that associates with RNA
polymerase and inactivates the elongation complex at a cytosine-rich termination site in the DNA.
Ø mRNA synthesis is catalyzed by ______ in eukaryotes. It then proceeds along the DNA template until a polyadenylation signal (polyA site) is encountered. At this point, the process of
termination of transcription is activated.
Ø There is _______ sequence in DNA that specifies
termination of transcription.
Ø As the polymerase proceeds pas the polyA site, the nascent mRNA is released by an _________ associated with the carboxy terminal end of polymerase II.
Ø RNA synthesized beyond the site trails out of the
polymerase and is bound by another __________
that begins to degrade the RNA 5’ to 3’ toward the
RNA polymerase.
Ø When the exonuclease catches up with the
polymerase, transcription ______.
Termination
```
Rho
polymerase II
no specific
endonuclease
exonuclease
stops
```
95
Give its template and product of this enzyme:
```
E.coli RNA polymerase II
RNA polymerase I
RNA polymerase II
RNA polymerase III
Mitochondrial RNA polymerase
Mammalian DNA polymerase α
HCV RNA polymerase
Dengue virus RNA polymerase
PolyA polymerase
```
```
DNA - mRNA
DNA - rRNA
DNA - mRNA
DNA - tRNA, snRNA
DNA - mRNA
DNA - primers
RNA - viral genome
RNA - viral genome
none - PolyA tails
```
96
Types and Structures of RNA:
Ø Largest component of cellular RNA, comprising 80-
90% of the total cellular RNA. An important structural
and functional part of the _____, cellular
organelles where proteins are synthesized.
```
Ribosomal RNA (rRNA)
ribosomes
```
97
Ø Initial connection between information stored in DNA
and translation apparatus that will ultimately produce
the protein products responsible for the phenotype.
Ø Amount of particular mRNA is related to the
requirement for its final product.
Messenger RNA (mRNA)
98
________– some messages are transcribed constantly and are relatively abundant in the cell
Constitutive transcription
99
__________– transcribed only at certain times during the cell cycle or under particular conditions
Inducible or regulatory transcription
100
__________ – most messengers carry a sequence of polyadenylic acid at the _____, a polyA tail. It is not coded in genomic DNA, but added to the RNA after synthesis of the pre-mRNA.
Polyadenylation
| 3’ terminus
101
____________ – enzyme responsible for
| adding the adenines to the end of the transcript.
Polyadenylate polymerase
102
__________– eukaryotic mRNA is blocked at the ______ by a 5’-5’ pyrophosphate bridge to a methylated guanosine. The structure is called a cap. It confers a protective function and serves as a recognition signal for the translational apparatus.
Capping
| 5’ terminus
103
________ – prokaryotic structural genes contain
____ lengths of open reading frame, sequences that code for amino acids. In eukaryotic coding regions, they are interrupted with long stretches of noncoding DNA sequences called _____.
They are then spliced or removed, and the remaining
sequences that code for the protein product are ____.
Splicing
uninterrupted
introns
exons
104
_________ – modifies products of genes by alternate insertion of different exons.
Alternative splicing
105
Ø Functions in splicing in eukaryotes
Ø Stays in nucleus after its transcription by RNA
polymerase I or III
Ø Isolated from _____ and cervical carcinoma cell
lines
Small nuclear RNA (snRNA)
hepatoma
106
Ø Relatively short, single-stranded polynucleotides.
There is at least one ___ for each amino acid.
Ø Translation of information from nucleic acid to
protein requires reading of the mRNA by ______,
using adaptor molecules or tRNA.
Ø Variable loop is larger in longer tRNAs. Another
seven-base loop is larger in longer tRNAs
Transfer RNA
ribosomes
107
RNA POLYMERASES
• RNA synthesis is catalyzed by _________enzymes.
• One ___________ enzyme is responsible for
the synthesis of all types of RNA in the prokaryotic cell.
• Eukaryotes have three different RNA polymerase
enzymes. All of these enzymes are __________ RNA
polymerases; that is, they require a DNA template.
• __________RNA polymerases require and RNA
template.
RNA polymerase
multisubunit prokaryotic
DNA-dependent
RNA-dependent
108
__________ – degrade RNA in a manner similar to the
degradation of DNA by ________. They are
ubiquitous and stable that degrade all types of RNA. RNA
work is performed because they are very resistant to
inactivation.
Ribonucleases
deoxyribonucleases
109
__________– RNA synthesis and processing require the
| activity of helicases to catalyze the unwinding of doublestranded RNA. Some of them work exclusively on RNA.
RNA Helicases
110
Ø Products of transcription and translation of nucleic
acids.
Ø Polymers of amino acids. Each amino acid has
characteristic biochemical properties determined by
the nature of its amino acid side chain.
Ø The most abundant macromolecules in cells. The
collection of proteins encoded in all of an organism’s
DNA is a _____.
Ø The ultimate effect of the information stored and
delivered by the nucleic acid is manifested in
____.
Proteins
proteome
proteins
111
Ø Even if proteins are not being tested directly, they
manifest the _______ directed by the nucleic acid
information
phenotype
112
In order to interpret the results of nucleic acid
analysis accurately, it is important to understand the
______________ from DNA to
protein as dictated by the genetic code.
movement of genetic information
113
__________
Ø Grouped according to their _____ or tendency to
interact with water at ___.
Ø _____, ________ polar, _______ charged
polar, and ______ charged polar.
Ø Properties of amino acids that make up a protein
determine the ___ and biochemical nature of the
protein. A single protein can have separate domains
with different proteins.
Ø Synthesized in vivo by ______ enzymes so that
naturally occurring proteins are made of amino acids
of L-stereochemistry.
Ø The ionization can switch between the amino and
carboxyl groups, making the amino acids _______
at physiological pH.
Amino acids
```
polarity; pH 7
Nonpolar, uncharged, negatively, positively
shape
stereospecific
zwitterions
```
114
Primary structure
§ The sequence of amino acids in a protein
determines the nature and activity of that
protein.
§ Read by convention from the _________ to the _________end.
§ _____ changes in primary structure can alter
the activity of proteins dramatically, because
the amino acids must _____________to bring about protein structure and function.
amino terminal end; carboxy terminal
Minor
often cooperate with one another
115
Ø Secondary structure
§ Interactions between amino acid side chains
fold a protein into _________.
§ Some proteins, especially structural proteins,
consist almost entirely of _________
§ Globular proteins have varying amounts of
alpha helices and beta sheets.
§ These include ordered beta or beta-pleated
sheets and less-ordered alpha helices, or
random coils; alpha-helix and beta-sheet
structures in proteins.
predictable configurations
| alpha helices or beta sheets.
116
Tertiary structure
§ Secondary structures are further folded and
arranged into a ___________.
§ If a protein loses its tertiary structure, it is
______.
§ ______ in DNA that substitute different
amino acids in the primary structure can also
alter tertiary structure.
tertiary structure
denatured
Mutations
117
§ ________ – proteins denatured by heat or by
conformations forced on innocuous peptides.
Aggregations of prions-induced folded
proteins cause transmissible __________, such as ________and bovine spongiform encephalitis
or _______.
Prions
spongiform encephalopathies
Creutzfeldt-Jakob disease
mad cow disease
118
Quaternary structure
§ Combinatorial nature of protein function may
account for the __________ of higher
organisms without a concurrent increase in
gene number.
§ Monomer, dimer, trimer, tetramer,
oligomers.
genetic complexity
119
_________
Ø Ordered sequence of nucleotides on a chromosome
Ø Encodes a specific functional product
Ø Fundamental ____and functional unit of
inheritance
Ø First studied by tracking mutations that took away
their function and observing the resulting phenotype
Ø Part of a chromosome responsible for the phenotype
affected by _____
Ø Contains structural and regulatory sequences
important for the regulated expression of the gene
Gene
physical
mutation
120
Ø ___________ – dictionary to translate the 4-
nucleotide sequence information in DNA to the 20-
amino acid sequence information in proteins
Genetic Code
121
TRANSLATION
After transcription of the sequence information in DNA to
RNA, the transcribed sequence must be transferred into
_______
• Through the genetic code, a specific nucleic acid sequence
is translated to an amino acid sequence and, ultimately, to
a ______.
• Takes place on ____, __________ particles
first observed by electron microscopy of animal cells.
proteins.
phenotype
ribosomes
ribonucleoprotein
122
_______ – molecular factor that can recognize components of
| both nucleic acid and protein sequences.
tRNA
123
TRANSLATION
In contrast to the analysis of phenotypic traits
(microscopic and colonial morphologies, enzyme or
pigment production, carbohydrate fermentation
patterns), the analyte for molecular testing is the _______,
transcriptome, or ______ of the microorganism.
genome
| proteome
124
Bacteria, fungi, and parasites have _____ genomes,
whereas viruses can have ______genomes. Prions,
which cause transmissible encephalopathies such as
Creutzfeldt-Jakob disease, consist only of _________
DNA
DNA or RNA
protein
