Combo with "Genetics Ch 5 Genetic Linkage and Mapping in Eukaryotes" and 14 others
parental ditypes (PD)
-Assuming no crossover between the homologs, ascospores produced by one of these arrangements contain the same alleles as the original parent strains -also 2-strand double crossover [no recombination]
second-division segregation pattern
-In a heterozygous meiocyte, if a crossover occurs between the gene and the centromere, the alleles segregate at meiosis II when sister chromatids segregate each ascus contains 4 parentals and 4 recombinant spores
In lab studies, the ascospores can be removed from the ascus and cultured to determine their genotypes; this process is called-
tetrad analysis
If a single crossover occurs between either of the homolog pairs, a _______ is produced, containing two parental spores and two nonparental spores
tetratype (TT)
Yeast artificial chromosome
The first artificial chromosomes developed:
odds ratio
-numerator=likelihood that distribution of genotypes and phenotypes in pedigree is produced by genetic linkage -denominator=chance that it is produced by independent assortment
Creighton and McClintock
-obtained cytological evidence that recombination between the genes was accompanied by physical exchange between the homologs -fragment on one end of chromosome knob on other
What is a genetic screen?
A procedure where a population is mutagenized and the progeny are examined for mutant phenotypes
Multiple cloning sites
A vector DNA sequence containing several unique restriction enzyme target sequences facilitating cloning of inserted DNA fragments
produce a complimentary strand to the target molecule and detect which bases are addded at each step. Millions of sequencing reactions can be run simultaneously, (high throughput) the DNA to be sequenced does not need to be known prior to sequencing. can sequence samples found in trace amounts, can sequence DNA that is difficult to clone
Advantages of next generation/third generation sequencing (4):
DNA doesn't need to undergo cloning or PCR, it is faster, it is cheaper
Advantages of third generation sequencing over next generation sequencing (3):
Primer walking or shotgun sequencing
After cloning a DNA fragment, one of these two methods can be used to sequence the DNA fragment to know what the nucleotide bases are:
Yes
Can blunt end DNA molecules be recombined?
100-200 kb
Capacity of a BAC
250-2000 kb
Capacity of a YAC
Under 15 kb
Capacity of a plasmid for insert sequences:
Under 23 kb
Capacity of linear phage chromosomes for insert sequences:
YAC's
Do BAC's or YAC's carry a larger insert size?
syntenic genes
Genes located on the same chromosome -so close together that alleles cannot assort independently
______ suggested that nonparental allele combinations resulted from recombination between the X chromosomes of the heterozygous female parent
Morgan -X linked gene for eye/wing in Drosophila
Using molecular probes
How are DNA libraries screened for specific DNA sequences?
interference [I]
I=1-c -the double crossovers expected but not produced -negative interference produces more double crossovers than expected
Double digest
In restriction mapping, in order to determine which order of restriction sites is correct, this must be performed:
Transformation
Recombinant DNA molecules are introduced into the bacteria by the process known as:
Sticky end
Short single-stranded overhangs created by the cleavage of DNA by specific restriction endonucleases, which can potentially base pair with complementary single stranded sequences
Shotgun sequencing
The sequencing method that relies on redundant sequencing of fragmented DNA in the hope that all regions will be sequenced at least a few times
Primer walking
The sequencing method that relies on the successive synthesis of primers based on the progressive attainment of new sequence information
Recombinant DNA technology
The set of techniques developed for amplifying, maintaining, and manipulating specific DNA sequences
Those derived from the genomic DNA of an organism (genomic lib), and those derived from mRNA
The two types of DNA libraries:
Genomic library
The type of DNA library consisting of genomic DNA
Complementary DNA library (cDNA library)
The type of DNA library consisting of mRNA
cDNA library
The type of DNA library that represents a subset of all the genes in the genome; represents only the expressed genes
Forward genetic analysis
The type of genetic analysis that starts with a genetic screen to identify specific phenotypic abnormalities in a population of organisms that have been mutagenized.
LacZ gene
What feature of the plasmid vector selects for only those bacteria that contain a recombinant plasmid and selects against nonrecombinant plasmids?
Add a Poly-dt primer for reverse transcriptase to use
What is the first step in making a cDNA library from mRNA?
Selects for only bacteria containing the plasmid
What is the purpose of a selectable marker in a plasmid vector?
Can only read about 1000 bp and most DNA regions of interest are much larger than this
Why can dideoxy sequencing not be used to sequence many DNA regions of interest?
transcription factors
a broad category of proteins that influence the ability of RNA polymerase to transcribe DNA into RNA.
true or false: The two parental types are approximately equal in frequency, as are the two recombinant types
TRUE
true or false: homologous crossover occurs sometimes during mitosis
TRUE
ordered ascus
arrangement of daughter spores reflects the orientation of alleles carried by each chromatid in metaphase I
linkage disequilibrium
certain haplotypes are maintained in a population, usually due to natural selection
_________ can be used to determine gene order
double recombinants, or double crossover -only one allele is different; this is the middle gene -list in each of 3 possible orders
Intragenic recombination
infrequent event in which crossing over occurs within a gene
tetratypes are [more/less] frequent than PD
less frequent -->NPD are even less frequent
The ascospores produced by the second arrangement will contain alleles in different combinations from the original parent strains, called a ______
nonparental ditype (NPD) -forming only when 4-strand double crossovers take place
coefficient of coincidence [c]=
observed double crossovers/expected double crossovers
GTF (general transcription factor)
one of several proteins that are necessary for basal transcription at the core promotor.
mapping function [based on Poisson distribution]
-does not account for interference -can be used to correlate the frequency of crossover with the actual number of crossover events
what crossover event is least frequent in 3-point test-cross?
-double crossover -both events must occur
an organism that exhibits complete linkage [rare]
-drosophila males have no crossing over
lod score
-logarithm of the odds ratio -determines theta value -sum all lod scores for each theta value to get Zmax -higher than 3.0=linkage at theta value -less than -2.0=significant evidence against genetic linkage -between -2.0 and 3.0 are inconclusive
when calculating recombination frequencies, for the larger distance, the double crossover progeny are-
-added twice because each represents 2 crossovers between them
three-point test-cross analysis
-allows you to map 3 linked genes simultaneously -
Genetic linkage
-can be quantified to map the positions of genes on chromosomes
Zmax
-corresponds to the recombination frequency that is most likely to be correct
theta value
-range 0-.50
upper limit to recombinant gametes-single crossovers
-single crossover produces 50% recombinant gametes and 50% parental -all 4 possible single crossovers result in same recombinant gametes
haplotype
-specific array of alleles in a set of linked genes on a single chromosome -these alleles tend to be passed on together during meiosis -may consist of any combination of linked genes with genetic variation
allelic phase
-the arrangement of alleles of linked genes on parental chromosomes -obstacle for mapping human genes -can be determined when disease-causing allele segregates with known genetic marker
why are genes that are closer together less likely to recombine?
-the closer the genes are, the less likely a recombination nodule will form between them
map units (m.u.) vs centiMorgan (cM) vs recombination frequency
1 m.u. = 1 cM = 1% recombination
Blunt ends
5' or 3' ends of double stranded DNA lacking any single stranded overhangs
what is recombination limit? why?
50% -Once genes are far enough apart to produce recombination frequencies of 50%, the pattern of segregation is that expected of independent assortment
Steps of a forward genetic screen
:Choose an organism, choose a mutagen, mutagenize the organism, screen for phenotypic defects, identify the gene sequence responsible for the mutation, analyse the molecular function of the gene
Vector
A carrier fragment of DNA with attributes that will allow amplification in a biological system
DNA library
A collection of cloned fragments of DNA (single source
DNA clone
A fragment of DNA that is inserted into a vector
Bacteria artificial chromosome
Cloning vector used in bacteria that utilizes the F plasmid origin of replication; can accept DNA inserts up to 500 kb
Yeast artificial chromosome
Cloning vector used in yeast that utilizes an endogenous yeast origin of replication, centromere and telomere; can accept DNA inserts in excess of 1 megabase
crossing over occurs at a ____ rate between genes that are further apart [lower/higher]
Crossing over occurs at a higher rate between genes that are farther apart, and a lower rate between genes that are closer together -higher rate = higher frequency
Mutagenesis
The process by which an organism is treated with a mutagen to create mutations throughout the genome
tetrad
The four ascospores in an ascus
Recombinant clone
The joining together of a cloning vector and donor DNA fragment produces a:
Molecular cloning
The method for DNA sequence amplification that does not require prior knowledge of the DNA sequence
PCR
The method that requires some prior knowledge of a DNA sequence that needs to be amplified
Within the lacZ gene
The multiple cloning site is often located here in a bacterial plasmid:
Bacteria artificial chromosome
The preferred artificial chromosome cloning vector
YAC
This cloning vector has the highest capacity
BAC
This cloning vector has the second highest capacity
Nonrecombinant DNA
This is created when the sticky ends of rejoin rather than incorporating a donor insert
Antibiotic resistance like ampicillan resistance
This is often the selectable marker in a plasmid vector:
Recombinant DNA technology
This provides the tools for proceeding from mutant phenotype to DNA sequence, so that alleleic differences can be examined at the molecular level
Restriction map
This shows the numbers and relative positions of target sites for restriction enzymes of a DNA molecule
Molecular cloning or PCR
To reproduce large amounts of individual pieces of DNA created by restriction enzymes, these two different processes can be used:
regulatory transcription factor
a protein or protein complex that binds to a regulatory element and influences the rate of transcription via RNA polymerase.
regulatory element (control element)
a sequence of DNA (or possibly RNA) that binds a regulatory protein and thereby influences gene expression. Bacterial operator sites and eukaryotic enhancers and silencers are examples.
what can affect recombination frequency?
age, environment, and sex -dietary levels of metal ions can affect crossover in flies
The diploid cells undergo meiosis to produce four haploid ______ in a sac called an _____
four haploid ASCOSPORES in a sac called an ASCUS
linkage equilibrium
genotype for a chromosome at one gene is expected to be independent of its genotype for other genes
Homologs that do not reshuffle alleles under study are called-
parental chromosomes or nonrecombinant chromosomes
Genetic linkage mapping-
plots the positions of genes on chromosomes
upper limit: Three-strand double crossovers
produce 50% parental and 50% recombinant gametes
upper limit: Two-strand double crossovers
produce ONLY PARENTAL gametes
upper limit: Four-strand double crossovers
produce all recombinant gametes -recombination LIMIT is 50%
crossing over occurs during-
prophase I of meiosis
r [recombination frequency]=
r= # recombinant gametes / total # meioses
recombination frequency=
r=[.5TT]+[NPD]/total tetrads -only half TT spores are recombinant
Alleles of syntenic genes can be reshuffled when crossing over occurs between homologs to produce ______________
recombinant chromosomes
gene regulation
the phenomenon in which the level of gene expression can vary under different conditions.
