Unit 6: Genetics

Genes

a heritable factor that consists of a length of DNA and influences a specific characteristic

Locus

address of the gene, allows for genes to be isolated, many genes can share the same loci

Alleles

alternative forms of a gene, caused by a difference in sequence of base pairs, codes for a specific version of a trait

alleles caused by _______

mutations: if a mutation is not lethal, it results in a new allele and will be passed down

Silent mutation

no new allele is formed, neutral

Nonsense mutation

likely harmful, often causes stop codon too soon

Missense mutation

new allele is formed, changes amino acid sequence, can be beneficial neutral or harmful

Sickle cell anemia base substitution mutation

base substitution on 6th codon for beta chain of hemoglobin, amino acid changes GAG to GUG and hence glutamic acid to valine (GLU → VAL)

Sickle cell anemia change in RBC structure

forms insoluble fibrous strands, changing RBC to sickle shape

Sickle cell anemia PROS

PROS: if 2 copies are inherited, very resistant to malaria since the parasite feeds on nutrition from healthy RBCs (especially K). If only one copy is inherited some sickle RBCs are still produced so individuals are still somewhat resistant

Sickle cell anemia CONS

CONS: cannot carry O₂ well so affected individuals is often tired, can clot in capillaries (recall only one cell diametre), and sickle RBCs destroyed more rapidly, causing anemia.

Genome

total genetic information of a cell, includes genes and non-coding DNA sequences

The Sanger Technique method for working out complete genome

1. DNA sample chopped into fragments of which copies are made, primer sequence added to help start the process

2. DNA polymerase adds free nucleotides, some of which are fluorescently marked ddNTPs, elongation stops when one of these are reached (they do not contain a 3’ hydroxyl)

3. gel electrophoresis used to order DNA strands long to short

4. laser activates fluorescent markers with a sensor attached to a computer

5. computer matches up sequenced code fragments

Applications of human genome project

• understand diseases that have a genetic basis

  • Can lead to genetic screening, using specific gene probes to detect sufferers and carriers

• genes can be used as ‘instructions’ to synthesize proteins in a lab that can be used as medicine

• Insights into origins, evolution, migratory patterns of humans

Prokaryote DNA

• genophore: single chromosome in a ring shape in nucleoid region

• plasmids: small DNA rings, not essential to life functions but contribute to evolution of thee cells within a generation via horizontal gene transfer, also reproduce independently of chromosomal DNA

• DNA is not associated with histones

Nucleosome

DNA strand wrapped around 8 histones, secured by a single H1 histone

Diploid chromosome number

2n, where n is haploid number, in humans 23 pairs of homologous chromosomes, 46 total

Haploid chromosome number

number of chromosomes in gametes, no homologous pairs

Karyogram

photo representation of chromosomes of an organism arranged by shape and size: decreasing length except sex chromosomes always pair 23. Taken at mitotic metaphase

Karyotype

specific number and appearance of chromosomes of an organism, represented by a karyogram

Sex chromosomes aka

heterosomes

Sex chromosomes in F vs M

F: XX M: XY

Homologous in females so sperm always determines sex

Cairns technique - autoradiography

• cells grown in solution containing radioactive T nucleotide, so incorporated into chromosomal DNA

• chromosome isolated onto photographic surface

• Surface then immersed into a radioactively sensitive emulsion

• Ag⁺ in the solution are converted to insoluble metal grains

• Chromosomal DNA can then be visualized with an electron microscope from black dots of silver grains

Meiosis

cell division with the purpose of creating haploid gametes, requires two cell divisions to reduce 2n to n

Crossing over/recombination

2 chromosomes ‘cross over’ and form a tetrad/bivalent, exchanging genetic material (alleles) where they overlap (chiasma)

interkinesis

2nd growth phase between meiosis 1 and 2 in which DNA is not replicated, does not always occur

random assortment

• during metaphase 1, homologous chromosomes line up at equator as bivalents in one of two arrangements (can have maternal/paternal copy either left or right)

• gametes differ depending on whether they got maternal or paternal chromosome

• occurs for each homologous pair so number of possible gamete combinations is 2^n where n is number of homologous pairs

spermatogenesis

Process of sperm cell production in the testes through mitosis and meiosis, resulting in the formation of mature sperm cells.

oogenesis

production of female egg gamete, only one produced per somatic cell, other 3 gametes become polar bodies and feed nutrients to one remaining gamete

nondisjunction

chromosomes failing to separate correctly, results in aneuploidy

Aneuploidy

having an irregular number of chromosomes in gametes

nondisjunction in meiosis 1 leads to _ affected gametes

4, two have + 1 chromosome, two have - 1 chromosome

nondisjunction in meiosis 2 leads to _ affected daughter gametes

2, one + 1 chromosome, one - 1 chromosome

Dominant alleles

are expressed if either heterozygous or homozygous

Recessive alleles

can only be expressed if homozygous (ie. no dominant allele present)

Genotype

an organisms set of genetic material (homologous pairs), represented by two letters when discussing a specific gene

Phenotype

the observable traits and characteristics of an organism that result from the interaction of the genotype and environment

Mendel’s law of segregation

when gametes form during mitosis, alleles are separated so each gamete carries only one allele for each gene

Mendel’s law of independent assortment

separation of alleles for one gene occurs independently of that of any other gene, however this does not hold true for genes located on the same chromosome (linked genes)

Mendel’s principle of dominance

recessive alleles will be masked by dominant alleles in most cases

Co-dominant alleles

Co-dominant alleles are two or more alleles that are equally expressed in a heterozygous individual. They are not dominant or recessive to each other

Incomplete dominance of alleles

When neither allele is dominant, and they blend together to create a new phenotype, it is called incomplete dominance.

Mono-hybrid cross

a genetic cross between two individuals that differ in only one trait

F₁ generation

first filial generation, results of first cross

F₂ generation

second filial generation, results of cross between products of F₁ cross

Antigens of RBCs

a surface glycoprotein on red blood cells that determines blood type

A and B alleles for blood type are ________

codominant, each modify the structure of the antigen to produce different variants

Type A possible blood donors

A or O

Type B possible blood donors

B or O

Type AB possible blood donors

A, B, AB, or O

Type O possible blood donors

only O

Type A: antibody produced

Anti-B

Type B: antibody produced

Anti-A

Type AB: antibody produced

neither anti-A or anti-B

Type O: antibody produced

both anti-A and anti-B

genetic diseases often cause by ______ allele

recessive, of autosomal genes. Because mutations that increase the difficulty of survival unlikely to be passed down since a parent is less likely to live a healthy life to breed. Mutations of recessive alleles can be masked and, hence can be passed on.

Cystic fibrosis

• autosomal recessive disorder

• caused by deletion mutation to CFTR gene on chromosome 7

• results in excess production of thick mucus → respiratory failure, pancreatic cysts

Huntingtons disease

• autosomal dominant disorder

• caused by mutation to Huntington (HTT) gene on chromosome 4

• causes repetition of CAG sequence, which is unstable, can cause protein to disfold and lead to neurodegeneration

• symptoms include spastic movement and dementia

Sex-linked genetic diseases

when the gene/allele for a disease is located on the X or Y 23rd chromosome pair.

• Most are X-linked, disproportionately affecting males if recessive

• If X-linked dominant, more common in F, either allele may be dominant and cause disease

Examples of X-linked sex-linked genetic disorders

• Red/green colour blindness

• hemophelia

• albinism

• muscular distrophy

Germline mutations

heritable

Somatic mutations

induced by an outside force in an individual’s lifespan

Recombinant DNA

DNA fragments composed of sequences originating from at least two different sources

Restriction endonucleases

• enzymes that are able to cleave double-stranded DNA into fragments at specific sequences

• isolated from and named after the bacteria they are extracted from, used in bacteria to digest foreign DNA

Recognition site

• a sequence of DNA that the restriction endonuclease is able to recognise and cleave

• at that point, phosphodiester bonds are broken by the enzyme via hydrolysis

Sticky ends on DNA fragments

• short single strands with complementary overhangs

• sticky end is often used to ensure a desired gene is inserted into the vector with the correct orientation and to prevent reannealing without the desired insert

Blunt end on DNA fragments

• the fragment ends are fully base paired with no overhang

DNA ligase

• reforms phosphodiester linkage if we want to put DNA back together after digestion

• T4 ligase from the T4 bacteriophage is often used to join blunt ends

PCR Use

• polymerase chain reaction

• an artificial method of replicating segments of DNA under laboratory conditions to amplify large quantities of a specific sequence of DNA from a minute sample

  • each reaction cycle doubles amount of DNA - standard sequence of 30 cycles results in >1billion copies (2^30)

PCR steps

• occurs in a thermal cycler and uses temperature variation to control replication

• 1) denaturation: DNA sample heated to separate it into 2 single strands

• 2) annealing: DNA primers attach to the 3’ end of the target sequence

• 3) elongation: a heat-tolerant DNA polymerase (Taq) binds to the primer and copies the strand

Gel electrophoresis

• separates and isolates proteins or DNA fragments based on mass/sizes

• Samples placed in a block of gel, an electric current is applied which causes the samples to move through the gel; smaller samples will move farther, larger samples will not move as far

Compare/contrast gel electrophoresis for DNA vs proteins

• DNA separation uses agarose

• Protein separation uses polyacrylamide

• For DNA, complementary radiolabelled probe used to transfer the separated sequences to a membrane and view using autoradiography (southern blotting)

• For proteins, separated proteins are transferred to a membrane and then target proteins are identified via staining with specific antibodies (western blotting)

DNA profiling

• satellite DNA fragments (long stretches of non-coding DNA made of repeating short tandem repeats -STRs) are separated by gel electrophoresis

• used to compare DNA profiles since each profile will have a different number of repeats at a given satellite DNA locus

• used for paternal testing and forensics

Process of creating a transgenic insulin-producing bacteria

• restriction endonuclease cuts/cleaves DNA segment from the human pancreas that produces insulin with sticky ends, which is then amplified by PCR

• plasmid removed from bacteria and used as a vector. It is cut at a recognition site with restriction endonuclease with sticky ends

• gene inserted into vector via complementary base pairing

• ligase combines them together to form a recombinant construct

• it is then inserted back into an E coli bacteria, which is now able to produce insulin

• it is then grown in a culture and the insulin can be extracted

Gene modification

transfer of genes between species, results in a transgenic organism

Genetic modification of crops: Bt corn

Bt corn incorporates a gene from the bacterium Bacillus thuringiensis which codes for the Bt toxin that kills corn feeding pests— corn bores

Pros and Cons of Bt corn

• Pros:

  • higher crop yield, more food for humans

  • less land needed for crops

  • less insecticide sprays used

• Cons:

  • Bt corn pollen may collect on milkweed and kill monarchs

  • leaves and stems remain toxic even after harvest, so can still kill detrivores

  • may spread to wild plants via cross-pollination

  • insects/pests of the corn may develop a resistance

Natural cloning in animals

• Binary fission - flatworms

• budding - hydra

• fragmentation - starfish

• parthenogenesis (embryo formed from unfertilized ova) - some fish, amphibians, reptiles, insects

• monozygotic twins -

Natural cloning in plants

• spores

• propogation

• runners/stolon

Somatic cell nuclear transfer

• cloned embryos can be produced using differentiated adult cells

• diploid somatic cell removed from adult and cultured

• unfertilized egg is removed from female adult and its haploid nucleus is removed to produce an enucleated egg

• this egg is fused with the nucleus from adult donor to make a diploid egg cell with donor’s DNA

• electrical current stimulates the egg to divide which leads to an embryo

Reproductive cloning vs Therapeutic cloning

• for reproductive cloning, the diploid egg produced by somatic cell nuclear transfer is implanted into the uterus of a surrogate

• for therapeutic cloning, the embryonic cells are used to differentiate to form specific tissues or organs for transplantation

Artificial cloning of embryos

embryonic cells are separated in a lab at the 8-cell stage, each resulting cell can be implanted into the uterus of the surrogate

Homologous chromosomes

• have the same shape and size, centromere position

• same genes at same loci, but can have different alleles

Chromosome arms

P arm = top arm, shorter

Q arm = bottom arm, longer

Haploid nuclei found in…

gametes, bacteria, fungi except when reproducing

Comparison of diploid numbers of different species

Parascaris equorum	Oryzo sativa	Homo sapiens	Pan troglodytes	Canis familiaris
Roundworm	Rice	Humans	Chimp	Dog
4	24	46	48	78
Pear	orzo with	homosexual	pan fried	dog

Comparison of genome size in different species

T2 phage virus	Escheria coli	Drosophila melanogaster	Homo sapiens	Paris japonica
170,000 bp	4.6 million bp	130 million bp	3.2 billion bp	150 billion bp
Virus	Bacteria	Fruit fly	Human	Canopy plant
Two f*gs	cheri and	Droso	are homosexuals	in Paris

General genome size statements

• virus and bacteria usually a smaller genome size

• eukaryotes > prokaryotes

• plant genomes vary greatly due to capacity to self fertilize and become polyploid

Sex chromosomes determination of sex characteristics

• SRY gene on the Y chromosome develops male sex characteristics

• female sex organs develop in its absence

Chromosome replication before meiosis

• suggests meiosis evolved from mitosis

• if not duplicated there would be no need for meiosis 2

• duplication increases potential for genetic recombination so more diversity

Maternal age and non-disjunction

• maternal age and incidence of non-disjunction is correlated

• higher chance of chromosomal errors in offspring if nondisjunction occurs in A1 not A2

obtaining cells for karyotype analysis (general)

• Chorionic villus sampling or amniocentesis

• karyotyping - chromosomes are arranged and visualized for analysis - gender or chromosomal abnormalities

• Cells harvested from foetus then chemically induced to undergo cell division so chromosomes are visible, then stained and photographed

Chorionic villus sampling

• sample of chorionic villus (placental tissue) harvested via a tube inserted through the cervix

• ~1% chance of miscarriage at 11 weeks

Amniocentesis

• amniotic fluid tested, extracted with a needle

• ~0.5% chance of miscarriage at 16 weeks

Hemizygous

contains only one allele eg. for a gene located on the X chromosome for a male

Codominant genetic disorders

• if a genetic disease is caused by co-dominant alleles, it will also only require one copy of the faulty allele to occur

  • but heterozygous individuals will have milder symptoms due to the moderating influence of a normal allele

• this is seen in sickle cell anemia

Morgan’s discovery of non-Mendelian ratios in Drosophila

• Studied fruit flys, phenotypes of vestigial vs normal wings, red vs white eyes

• Found red eyes = x-linked

• also determined linked genes don’t segregate independently

• and that they can be uncoupled by recombination to create alternative phenotypic ratios

Frequence of recombination with lined genes depends on…

the distance between them; recombination occurs more frequently if they are farther apart; more possible locations where a chiasma could form between the genes

Test cross

crossing an organism with homozygous recessive, can be used to identify if genes or linked; ratios will not resemble 9:3:3:1

Polygenic characteristics variation

• polygenic characteristics are traits coded for by multiple different genes

• they show continuous variation, the phenotypes of individuals exist along a spectrum

  • the more genes contributing, the more possible phenotypes (not just “yes” or “no”)

• Resembles a gaussian normal curve eventually