Genetics exam 3

types of chromosomal alterations

• duplications

  • partial, chromosomal, genomic

• deletions

• rearrangements (translocation, inversion)

chromosomal duplications

duplicating entire chromosomes, increases number of genes, we’re overexpressing those genes

polyploidy (every chromosome has too many) vs aneuploidy (one chromosome has too many)

segmental duplication

makes localized copies of genes

• tandem duplication: copy inserts adjacent to the original, same or reversed gene order

• dispersed duplication: copy inserts somewhere else on the same chromosome or on another chromosome (different regulatory neighborhood, can end up in eu/heterochromatin, etc.)

results in overexpression/failure of dosage compensation for that particular gene

can cause developmental conditions

aneuploidy

duplication of individual chromosomes

unbalanced karyotype is typically problematic

• caused by:

• nondisjunction in meiosis

• deletion/insufficiency of centromere

• mitotic errors during zygote formation

  • results in mosaic aneuploidy (some of the cell type is mutated, some isn’t, there’s usually enough normal ones that it doesn’t matter)

usually do not develop in humans, except for XY chromosomes due to X inactivation

nullisomy

2n-2 chromosomes, missing a homologous pair

monosomy

2n-1 chromosomes, missing one homologue

trisomy

2n+1 chromosomes, extra homologue

tetrasomy

2n+2 chromosomes, two extra homologues of a pair

nonlethal human trisomies

• Down’s Syndrome: Trisomy 21

• Patau Syndrome: Trisomy 13

• Edward Syndrome: Trisomy 18

All cause developmental abnormalities, Down’s is the least severe

Polyploidy

Duplication of the entire genome

can be caused by failure to separate in meiosis II, or two sperm fertilize the same egg

Many species will survive, or a lot of hybrids are triploidy, but odd numbers of chromosomes can’t reproduce after

any amount is usually fine for clonal species

autopolyploidy

extra set(s) of chromosomes from the same species

lethal in humans

for even numbers, all homologues are still matching

even numbers are fertile, odd numbers are infertile

form multivalents in meiosis, complex patterns of linkage and crossing over

more than two alleles possible for one gene

occurs from: failure of spindle separation/cytokinesis of the egg

diploidization

genome is polyploid, but it gets fixed, bivalence reestablished, often tetraploid

the duplicated genes mutate a lot and because there are still other functioning copies, the mutations don’t kill evolutionarily

the duplicated pairs end up matching up and forming new orthologous pairs (2n = 8 becomes 4n = 16 becomes 2n = 16)

allopolyploidal

duplication of sets of chromosomes in hybrids, basically combine two complete genomes in one cell

interspecific chromosomes don’t pair in meiosis, “multidiploid”

more genetic diversity, very adaptable

chromosomal deletions

part of the DNA breaks off, the DNA gets repaired without it

causes developmental disabilities

haploinsufficiency

• loss of one allele due to a chromosomal deletion when the gene needs two functional copies to produce a typical phenotype

• pseudodominance: recessive genes are dominant by default

• unbalanced gene expression

chromosomal inversions

take part of the DNA out, reverse it, put it back in (balanced rearrangement)

have all the same genes in a different order

paracentric: not on centromere

• during crossover in meiosis, have to twist to match up the homologous genes, creates inversion loops with dicentric bridges

• results in loss of certain chunks that don’t have a centromere, makes nonviable gametes

• nonrecombinants are fine

pericentric: includes centromere

• they don’t lose chunks, but the genes aren’t matched and recombinants don’t have the right sets of genes, end up with two of one gene and none of another, nonviable gametes

** if inversion if homozygous (both homologues do it), crossing over is fine, no inversion loops

large inversions have higher potential for crossover/recombinants

smaller inversions are often fine because they don’t recombine

translocations

non-homologous chromosomes swap segments

reciprocal: two chromosomes trade segments

non-reciprocal: one chromosome donates segments

robertsonian translocation: occurs near centromere of acrocentric chromosome (right near the edge), not a lot of genes. then, the two long segments join, and the two smaller segments join, so you have one long chromosome

balanced until meiosis occurs

determination

cells develop a cell fate determination, they lose their ability to become other types of cells

syncytium

one “cell” where the fertilized egg proliferates rapidly but does not undergo cytokinesis: a bunch of nuclei without separating cell membranes, allows very fast growth

occurs in some fish and flies as the early stage of development

the nuclei migrate to the edges and then form membranes

primordial germ cells

cells that are used to make gametes are isolated very early in development before any damage can occur to DNA

somatic cells

non-gamete producing cells

maternal factors

when mom produces an egg, she puts in a bunch of RNA and proteins that drive initial growth and development of the fertilized egg, promote cell growth and replication

after cdc25 used up, then the embryo itself takes over

mitosis at this point is all there is, no interphase, the cells aren’t doing anything

MZT

maternal zygotic transition, the zygote genome takes over regulation of development of the embryo and uses its own transcription factors and genes to drive growth of the organism

introduces interphase so gene regulation and protein creation can occur

polarity

dorsal/ventral, anterior/posterior patterning of the embryo, one of the first thing that develops in humans

driven by maternal factors

morphogens

secreted ligands that form gradients in the embryo, bind to cells, create polarity that determines cell fates

often different things happen at high C, low C, or sometimes middle C

genes are regulated by the morphogens, they express specific genes which turns them into certain cell types

homeotic genes

determine where in the embryo things are positioned, allows certain genes to be expressed in the right position

master regulators of the developmental program

made of Hox genes related to anterior/posterior patterning, encode transcription factors of certain segments of the embryo with homeobox motifs (helix turn helix, DNA binding motif)

role of apoptosis in development

we form structures by removing tissue, like carving them out

removal of early tissue, like the webbing between fingers

cellular immunity

T-cells, macrophages, complete cells that remove toxins and antigens

humoral immunity

secreted antibodies, not live cells (produced by B cells, immune system selects B cells with desired antibodies)

tag antigen for the immune cells to come and destroy

how do genetics impact immune system

we have DNA sequences that encode the constant regions of antibodies, and several different ones that encode different variable and joining regions. The DNA recombines to create groups of these three, to form different antibodies which can then be expressed over and over again. Also some splicing. every new B cell does this in a different way

extrachromosomal DNA

DNA pieces in bacteria that carry non-essential but helpful genes, allows them to pass genes and amplify genes with multiple copies without modifying the genome, carried in plasmids

episomes

plasmids which can integrate into the main genome

F factor

plasmid, determines mating type

made of selfish DNA: only exists to replicate

Contains machinery to conjugate with other cells, without incorporating into the genome just makes other cells F+ by passing the F factor

Hfr cells (high frequency of recombination) incorporate F factor into the genome

Then, when the F factor is conjugated to another cell, it takes some of the chromosomal DNA with it, this doesn’t make the recipient F+

conjugation

bacteria directly exchange DNA

• F+ cell recruits F- cell

• Membranes fuse, form a cytoplasmic bridge

• Part of donor gene gets cut out of donor genome and threaded into the recipient (recopied from donor’s other copy) and inserted into the homologous gene location

• crossover can occur with the two homologues

• the rest of the DNA is degraded

transformation

bacteria incorporate foreign DNA from the environment (either from another cell or just somewhere else) (or eukaryotes can do this too)

homologous recombination

change in bacterial phenotype

happens to competent cells: holes in the membrane (can do this in the lab, or some proteins will do it for transport purposes)

transduction

incorporation of viral DNA into the bacteria (or other types of cells)

virus infects cell, injects DNA, then the bacteria make more virus

sometimes the viruses pick up bacterial DNA and then insert it into other bacteria, leading to recombination

viral genetics

very small RNA or DNA genome

deliver their genetic material into another cell, take over the mechanism of the cell, sometimes genetic material is integrated into genome

makes cell make more virus

bacteriophage

viruses that infect bacteria

Lytic cycle

phage injects material into bacteria, host DNA is destroyed, host replicates phage DNA and translates proteins, mature phage is created, host cell is lysed and the mature phage released

Lysogenic cycle

phage injects genetic material, gets incorporated into bacterial genome, the offspring are all infected until some point at which the prophage (phage genes) start expressing, and enter the lytic cycle

can create a bunch of virus all at once, can overwhelm an organism’s immune system

balanced vs unbalanced rearrangements

balanced: no change in amount of genetic information on single chromosome (e.g. inversions)

unbalanced: some loss or gain in genetic information (relative to wildtype) (e.g. insertions/deletions, duplications)

cyclin/cdk

proteins that drive first 14 cycles of mitosis (cyclin & cyclin dependent kinases)

cdc25

phosphatase, gets degraded with each division, activates cdks and allows it with cyclin to drive mitosis, as it degrades they’re not able to do this anymore

describe dorsal/ventral process

dorsal mRNA makes dorsal protein, theres a gradient of cactus, low cactus means dorsal goes to the nucleus and that cell becomes ventral. high cactus blocks dorsal, those cells become dorsal

anterior/posterior process

• maternal protein bicoid is high at future anterior

  • promotes hunchback expression, head/thorax

• maternal protein nanos is high at future posterior

  • inhibits hunchback expression, defaults to abdomen

gel electrophoresis explanation

DNA is neg. charged, moves in an electrical field, gel impedes movement by size, DNA tagged with fluorescent dye

creates banding patterns of DNA fragments based on size

capillary electrophoresis is an automated version of this

how can we clone individual genes

isolate gene with PCR, use a restriction enzyme to cut that DNA and a bacterial plasmid, ligate the two together and infect a bacteria, bacteria duplicates the gene

PCR

polymerase chain reaction

• make DNA over and over again

• use heat to unwind DNA

• use DNA pol III to start replication, providing synthetic primers for the specific gene

• makes many copies

• uses a bacterial DNA pol that is resistant to heat/doesn’t denature when hot

• has a lot of room for error, we don’t have all the usual proofreading mechanisms

• can make some then put it in bacteria to make more with more accuracy

restriction enzyme digest

• bacteria have a restriction enzyme that cuts DNA at a specific sequence (palindrome, same forwards and backwards)

• use it to cut DNA, leaving an overhang (one piece is longer), can then stick together with other strands

• use these to have access sites on plasmid vectors, put palindromes in plasmid, we know the enzyme will cut it, and put DNA cut the same way in there

• plasmid also has an ABX resistance gene to kill off the bacteria without the plasmid/inserted gene

• used for fingerprinting and genetic testing

• HindIII cuts between 2 As on the 5’ end

• EcoRi cuts between a GA on the 5’ end

cosmid

plasmid/phage hybrid, injects DNA into bacteria that then gets replicated like a plasmid

BAC

bacteria artificial chromosome

RFLP

restriction fragment length polymorphisms

• cut DNA with various restriction enzymes at the natural palindromes

• different people have different palindromes in different places, banding patterns of the fragments leave DNA fingerprints

• use gel bands

microsatellite markers

repeat sequences in DNA that can be tracked based on number of repeats/allele length

cloning vectors

ways to put human or other genes into bacteria to clone them

• plasmid vectors (10 kb insert)

• viral DNA (23 kb insert)

• cosmid vectors (44 kb insert)

• BACs (300 kb insert)

allele detection

use PCR to detect different mutants of a gene for genetic testing

sanger sequencing

PCR with a small amount of fluorescent nucleotides, halts DNA replication

PCR makes a bunch of copies stopped in different places with a dye on the end

by reading the colored peaks we can see at which position has what nucleotides, sequence the gene

genomic libraries

entire genome collected, fragmented, put into vectors and then into bacteria and stored this way for further use/study

mutagenesis

change DNA sequence (random or directed)

things we need on an expression construct

promoter & operator, then shine delgarno sequence before the gene

terminator after the gene

elsewhere: ori, selectable marker like ABX resistance, repressor for an inducible protein gene (like if the protein would kill the bacteria)

for eukaryotes: instead of SDS, it’s the kozak sequence, we need polyA signal and 5’ and 3’ UTRs, some other selectable marker

GMOs

genetically modified organisms with genes from another organism inserted into theirs

cDNA libraries

made from total mRNA in a cell, reverse transcribed into DNA, lack introns so only get protein coding genes

how can we tag expression constructs to study protein expression in the cell

• expression constructs can code proteins that include something like GFP (DNA for the protein and then the tagged protein right after), or link a promoter of interest to GFP

• then visualize protein interactions or cell transport dynamics

horizontal gene transfer

DNA from one species gets incorporated into the DNA of another species

transgenesis

move a gene from one organism to another

change phenotype in novel way, add/remove control elements, insert reporter gene (GFP, lacZ)

lipofection

put DNA in a ball of phospholipids, inserts into eukaryotes

adenovirus

viral DNA in cell but does not insert into genome, eventually recognized by immune system and destroyed, disease causing often

agrobacterium

virus like bacteria that can inject genetic material into plant cells

gene therapy

replace bad genes

• swap it with homologous recombination

• add an extra copy

• in mammals, usually with viral transduction

genomics

organization of genetic elements on chromosome

linkage maps

frequency of recombination between genes, rough distance from each other and position

physical maps

using DNA sequencing to actually see where genes are located on chromosomes, often correspond with linkage but not always, some areas are more or less likely to recombine for reasons more than distance

map based assembly

breaks DNA in long fragments, recognize known protein coding genes, organize based on linkage map, gives the relative order of things but doesn’t account for all non coding DNA

shotgun assembly

breaks DNA into smaller fragments, puts them together one by one using sequencing and recognizing short overlapping sequences between fragments

SNPs

single nucleotide polymorphisms

single nucleotides in areas that are highly conserved, but vary in one position between individuals, often have patterns (called haplotypes) used to tell people apart

these haplotypes often go together in populations

GWAS

genome wide association studies

SNP/haplotype markers associated with certain phenotypes or diseases, based on location of SNP allows us to identify relevant genes