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Micro Chap 10
Terms in this set (81)
methods for studying the entire genome of a microbe
genomics has been spurred by the development of _________________ _______ protocols
however, the increase of genomics has created several new needs such as improved _____ _______________ techniques
this method of DNA sequencing requires three steps:
1. cloning of gene fragment to be sequenced
2. DNA synthesis
dieoxy sequencing is also called _________
by placing __________________ (lack the free 3' OH group) into DNA synthesis mixture, the process is terminated with a distinct, labeled endpoint nucleotide
______ ________________ can separate the fragments of different length and detect which labeled nucleotide is on the end of each fragment, providing a sequence
automated methods using _________________ labels instead of radioactive laves are safer, cheaper, and easier
longer sequences are obtained by "___________ walking" or using repeated rounds of sequencing with primers complementary to the end of the last segment sequenced
next generation sequencing is called
-detects addition of a nucleotide to the end of a synthesized strand of DNA by production of light
-faster and cheaper than sanger method
-high-throughout pyrosequencing method
-DNA doesn't have to be cloned first, only fragmented
-method similar to 454 sequencing but attempts to sequence entire genome in one setup
-DNA fragments are sheared and sequenced
-software aligns sequences
-may need 10x total genome length to do successfully
analysis of large data sets of sequencing data
-annotation od genomes help researches identify _______ _______ ________ (ORFs)
-ORFs allow us to better determine the start and stop points for a given gene
open reading frame
the goal of __________ ______________ is to determine the function of genes and not just sequences
-the method of obtaining the library varies depending on the desire outcome
-generated by shearing the genome and cloning the fragments
collection of transcribed mRNA molecules in a cell
separation of RNA fragments by electrophoresis, followed by blot transfer and probing with labeled DNA fragments
method for examining transcriptional activity of all genes in a cell simultaneously
this technique is a miniature, automated reverse of norther blots
a library of the expressed mRNA molecules of a cell can be formed as a ______________ DNA (cDNA) library using reverse transcriptase
-probe DNA fragments are amplified by PCS and placed on a glass slide in a specific, known pattern
-total cell mRNA is converted to cDNA by reverse transcriptase, label with a fluorescent molecule and passed over the slide
-the more intensely a "spot" lights up,the more cDNA and mRNA is present
example of this method is comparison of mRNA expression profiles in Y. pests microbes vowing at flea or human body temperatures
RNA sequencing is performed by conversion of mRNA into cDNA by _________ _______________
has been used to successfully to study Saccharomyces cerevisiae
-the collection of expressed proteins in a cell
allows separation of proteins on a gel based on 2 things:
1. isoelectric points
pH where protein has no charge
separation of the gel, smaller fragments move faster
___________ __________________ can be used to determine amino acid sequences of portions of the polypeptides from 2D-PAGE
2 methods for determines 3D protein shape:
X-ray beam shot at crystallized protein
-diffraction pattern can be used to discern protein shape
-measure distance between atomic nuclei to discern protein shape
-can measure proteins in solution
-limited to proteins about 30 kDa
genes that are different from each other, yet rose from a duplication even
genes that have evolved from the same ancestor with the same function in different organisms
areas of genomes with significantly lower G and C content than the rest of the genome are likely ares where ____________ ____________ has occured
involves construction and analysis of gene libraries from DNA extracted directly from complex microbial communities
Term for the process in which a peptide fragment expressed from a plasmid is able to combine with a peptide fragment expressed from the chromosome to form a functional peptide.
b) Peptide binding
c) Reciprocal crossover
e) Genetic complementation
Which of these is contained in an expression vector but not in a regular cloning vector?
a) restriction enzyme site
b) antibiotic resistance gene
c) strong promoter
d) origin of replication
e) selectable marker
A his-tag or strep-tag is part of the cloning area in expression vectors. How do these sequences function?
a) To increase gene expression from the cloned DNA.
b) In regulation of expression of the cloned DNA.
c) As unique restriction enzyme sites to aid in cloning DNA.
d) Affinity sequences to aid in purification of the recombinant protein.
e) As signal sequences for the secretion of the recombinant protein.
Which of these is another term for the Sanger method of DNA sequencing?
a) chemical degradation method
b) dideoxy sequencing
c) restriction method
d) label method
e) gel method
The Sanger method of sequencing uses ALL of the following EXCEPT:
b) DNA polymerase.
c) a short oligo primer.
e) restriction enzymes.
The Sanger method of DNA sequencing has been automated and uses a(n):
a) thermal-stable DNA polymerase to allow for multiple rounds of DNA synthesis.
b) trideoxynucleotides in the synthesis reactions.
c) radiolabeled nucleotides for fragment detection.
d) agarose gel electrophoresis for fragment separation.
e) chemical digestion method for fragment generation.
What is an open-reading frame?
a) A non-translated region on the chromosome.
b) an unknown gene.
c) A gene that no longer has a function.
d) The protein encoding sequence of a gene.
e) The intron region of a gene.
Many scientists deposit their gene sequences in GenBank to make them publicly available to other scientist via the internet. Approximately how many base pairs have been deposited thus far?
c) 1 million
e) greater than 10 million
What is a transcriptome?
a) The coding region on the chromosome.
b) The coding region on the chromosome minus the introns.
c) The coding region on the chromosome plus the control regions.
d) The transcripts encoded for by the genes within a genome.
e) The mount of mRNA in the cell.
What was the Northern blot hybridization technique used to measure?
a) The transcriptional expression of a specific gene.
b) The translational expression of a specific gene.
c) The the frequency of transcription initiation.
d) The amount of DNA in a cell.
e) The rate of chromosomal replication.
Microarray analysis may be used to study:
a) gene expression in cells grown under different conditions.
b) protein expression in cells grown under different conditions.
c) rate of chromosomal replication in cells.
d) enzyme activity in cells.
e) growth rate of cells when grown under different conditions.
Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) separates proteins based on what property?
d) size and charge
e) size and hydrophobicity
Which of these can be used to determine protein structure?
a) amino acid sequencing
b) X-ray crystallography and NMR
c) polyacrylamide gel electrophoresis (PAGE)
d) denaturing gel electrophoresis
e) mass spectrometry
What is the isoelectric point of a protein?
a) The pH at which the protein has the greatest charge.
b) The pH at which a protein has a negative charge.
c) The pH at which a protein has no charge.
d) The proton potential of a protein in an acrylamide gel.
e) The proton potential of a protein in a cell.
What are paralogs?
a) Genes that arise from a duplication event in an organism.
b) Mutant genes in an organism.
c) Genes that are seldom expressed in an organism.
d) Genes that are expressed at a high level in an organism.
e) Genes that have no function in an organism.
Term for genes from different organisms that encode for proteins that carry out the same function but have different sequences.
Which of these is TRUE regarding horizontal gene transfer?
a) It does not result in permanent change.
b) It does not play a role in microbial evolution.
c) It does not result in any significant consequences for the organism involved.
d) It is always occurring in nature.
e) It does not alter the genome of the organism.
Suppose it is observed that a small section of a microorganism's chromosome varies significantly in GC content from the majority of the chromosome. What likely caused this?
a) spontaneous mutations
b) induced mutations
c) horizontal gene transfer
d) vertical gene transfer
e) site-directed mutagenesis
What is metagenomics?
a) A culture-dependent method that may be used to determine genome sequences.
b) A culture-dependent method that may be used to find new enzymes.
c) A culture-dependent method that may be used to study groups of microbes in an ecosystem.
d) A culture-independent method that may be used to document microbial community composition.
e) A culture -independent method that may be used for the isolation of different organisms.
Transformants are recombinant bacteria that have received new DNA via a bacteriophage.
An expression vector contains a strong promoter next to the cloning site.
Affinity tags often used in modern expression vectors to produce a fusion protein from the gene of interest, aids in recombinant protein purification.
The Sanger method of DNA sequencing requires an oligonucleotide primer.
An open-reading frame (ORF) is the non-translated part of a gene.
a transcriptome refers to all the proteins produced in a single cell.
The Southern blot uses a DNA probe to hybridize to DNA fragments that have been separated by electrophoresis.
DNA microarray technology allows a researcher to analyze gene expression in an organism cultured under different growth conditions.
The isoelectric point is the pH at which a protein has zero charge.
Paralogs are genes that arose from a duplication event in the cell.
Orthologs are genes that arose from a duplication event in the cell.
Horizontal gene transfer occurs when an organism acquires genetic information from another organism in its own generation.
Analysis of the genome sequence of an organism shows that an area of the genome contains stretch of several thousand base pairs that differ significantly in % G + C content from the rest of the genome. Most likely this DNA was originally obtained from another organism.
Metagenomics uses a culture-independent based genomic analysis of microbial communities to determine community composition.
recombinant DNA technology was greatly facilitated by the discovery of ___________ enzymes
the sanger method of sequencing used special nucleotides that are called ____
In analyzing a genome sequence, a computer program identifies the beginning and the end of a protein encoding sequence. This sequence is referred to as a(n) _______ _______ frame.
The complete complement of proteins in the cell that results from the translation of all the mRNA transcripts is called the ____________.
A technique used to examine all mRNA production in the cell simultaneously for any set of growth conditions is called a/an ________________.
The pH at which a protein's charge is completely neutral is the protein's ______________ point.
A gene in a cell undergoes duplication. This gene may evolve a novel function while the original gene encodes for a protein with normal function. These genes would be considered ____________.
Genes from different organisms that differ significantly in their sequence but encode for proteins that perform identical functions are called ________.
___________________ gene transfer occurs when an organism acquires a gene from another organism and incorporates the genetic information into its genome.
A culture-independent method that is used to study genomes from microbial communities in their natural environments is called _______________ .
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