BBT: 511 Genetics

Dr. Bipinraj

Significance of Genetics in Evolution and Breeding

Genetics : study of heredity and variation Genetics helps us to understand heredity and variation, thus mechanism of evolution. Breeding is the application of genetic principles of to get better variety

 Evolution: Is the genetic changes in successive generations. Evolution give rise to diversity and causes speciation Evolution occurs when there is a variation in the allelic frequency in the population Variation in allelic frequency can happen due to natural selection, genetic drift and mutation.

Industrial Melanism

 Applications of Evolution:
Artificial selection or selective breeding to get desired variety Medicine to control genetic disorder

 Plant breeding: It is the artificial breeding to

Increase quality and yield Increase resistance against
Draught, salinity, temperature Virus, fungus and bacteria

Increase tolerance against pesticide and herbicide

Classical breeding : relies on homologous recombination chromosomes to introduce desired variety Modern breeding : use molecular techniques to select or introduce desired traits

 Steps involved are

Creation of variation, Selection and Evaluation
controlled pollination, genetic engineering Marker assisted selection

Release Multiplication Distribution of the new variety

CHROMOSOME

 Chromosome is the orderly arrangement of DNA. In eukaryote, DNA double helix is bound to an octamer protein (HISTONE) to form a beadlike structure called as (NUCLEOSOME ) Nucleosomes are connected by linker DNA strand Nucleosomes are further coiled to form solenoid tubes

Solinoid tube

 Solinoid tubes now spirally arranged on a nonhistone protein scaffold and this scaffold further coils to form condensed chromosome. On condensed chromatin the region at which spindle fiber attaches is called as centromere On chromosome arm there are more condensed region called heterochomatin and less condensed region called as euchromatin

 Cetromere Arms
p (shorter) q (longer)

Telomere

 Types of Chromosomes
based on centromere
Telocentric : cetromere at the end Acrocentric : centromere at off center Metacentric : Centromere in the middle Holocentric : Whole chromosome act as centromere

Sex chromosome Autosomes

 Staining: Used for visualizing the chromosomes

aceto-orcein, acetocarmine, gentian violet, haematoxylin etc. Giemsa, Wright and Leishman

Applications:
Chromosome analysis
Number Morphology Length Position of centromere etc.

To study chromosome breakage

 Giemsa staining:
Procedure
Place slides in a Coplin jar or staining dish. Stain for 7 minutes. Rinse slides in two changes of distilled water. Air dry slides; mount them with a cover slip if desired.

 Chromosome banding:
Differential intensity pattern along the length of chromosome when stained are called as chromosome banding. This difference is due to the differences in the base component of DNA Based on the stain used, chromosome produce different banding pattern

 Uses of banding
Chromosome identification Identifying abnormalities of chromosome number Identifying translocations Identifying deletions, inversions or amplifications Comparisons of chromosome banding patterns can confirm evolutionary relationships between species

 Standard banding methods

Q-Banding: pattern created by treating fluorochrome quinacrine. Forms dark bands at AT rich regions (heterochomatid) producing G-Banding: pattern created by treating Giemsa stain. Forms dark bands at AT rich regions (heterochomatid) producing R-Banding : Reverse banding, produced by staining GC rich region by dyes such as chromomycin A3, olivomycin and mithramycin. C-Banding : Banding of non-coding constitutive heterochomotin by Giemsa staining. C-Banding is produced by treatment of chromatin with acidic and then basic solutions followed by staining with Giemsa.

 High resolution banding

To produce more detailed analysis of chromosome
Hy-banding can be produced for entire chromosome or a specific region

Methods:
Banding prophase or early metaphase chromosome By decreasing the degree of condensation by treating with chemicals such as Ethidium bromide, acridine orange and actinomycin D etc.

 Fluorescence in situ hybridization: it is technique used to detect the presence or absence of a specific DNA sequence or gene.

FISH

Karyotyping
Karyotyping is the process of pairing and ordering all the chromosomes of an organism, thus providing a genome-wide snapshot of an individual's chromosomes. Karyotypes are prepared using standardized staining procedures that reveal characteristic structural features for each chromosome. Applications:
To detect gross genetic changesanomalies involving several megabases or more of DNA. Karyotypes can reveal changes in chromosome number Careful analysis of karyotypes can also reveal more subtle structural changes, such as chromosomal deletion, duplications, translocations, or inversions. Karyotypes are becoming a source of diagnostic information for specific birth defects, genetic disorders, and even cancers.

 Preparation of Karyotyping
Prepared from mitotic cells that have been arrested in the metaphase or prometaphase Burst the cell and fix the nuclei on a glass slide Treat the fixed cell with stains Take the image of individual chromosome Arrange them according to the size Align them on a horizontal axis, keeping centromere on the axis. individual chromosomes are always dipicted with its p arm on top and q at the bottom

 Nomenclature:
Standard nomenclature of chromosome is done by the size of the chromosome, So in human, all non-sex chromosome arranged according to the size, and 22nd, the sex chromosome arranged at the end. They are again sub divided into 7 groups (A-G) based on the size and position of centromere.

Group Group A Group B Group C Group D

Chromoso me no. 1-3 4-5 6-12 13-15

Size and position of centromere Largest , Median centromere Large, Submedian centromere Medium, Submedian centromere Medium, Acrocentric centromere

Group E
Group F

16-18
19-20

Group G

21-22

Short, Median or submedian centromere Short, Median centromere Very short, Acrocentric centromere

Heredity
Genetics is based on hereditary variation
Continuous variation Discontinuous variation

Inheritance
Autosomal inheritance Sex-linked inheritance Cytoplasmic inheritance

 Autosomal dominant inheritance :

Patterns of inheritance

One mutated copy of the gene in each cell is sufficient for a person to be affected by an autosomal dominant disorder.

Eg: Huntington disease

Progressive brain disorder that causes uncontrolled movements, emotional problems, and loss of thinking ability Mutation in HTT gene : important in neuron function CAG repeats 10 35 in normal 36 120 in abnormal

Autosomal recessive
Two mutated copies of the gene are present in each cell when a person has an autosomal recessive disorder. Eg: Cystic fibrosis
Inherited disease of the mucus glands include progressive damage to the respiratory system and chronic digestive system problems Mutations in CFTR gene Impaired function of the chloride channels, preventing them from regulating the flow of chloride ions and water across cell membranes

X-linked dominant
Caused by dominant mutations in genes on the X chromosome Eg: Fragile X syndrome
This disease is the most common form of inherited mental retardation It is manifested cytologically by a fragile site in the X chromosome that results in breaks in vitro

X-linked recessive
Caused by recessive mutations in genes on the X chromosome Eg: Hemophilia
Due to the absence or malformation of factor VIII

 In codominant inheritance, two different versions (alleles) of a gene can be expressed, and each version makes a slightly different protein. Both alleles influence the genetic trait or determine the characteristics of the genetic condition.

Codominant

Eg: ABO blood group

Mitochondrial

Eg: Optic neuropathy

GENE INTERACTION
Influence of two or more non-allelic pairs of gene over a character
Epistasis

 Epistasis:
One gene override the expression of other gene
Expressed gene is called as epistatic Overridden gene is called as hypostatic Dominant Recessive Complementary Duplicate

9 C_P_ : 3 C_pp :3 ccP_ : 1 ccpp purple white

 Complementary gene action

Epistatic Gene Interaction

Enzyme C and enzyme P cooperate to make a product, therefore they complement one another
Enzyme C Enzyme P

Colorless precursor

Colorless intermediate

Purple pigment

Colorless precursor

Enzyme C

Colorless intermediate

Enzyme P

Purple pigment

genotype cc

Colorless precursor

Enzyme C

Colorless intermediate

Enzyme P

Purple pigment

genotype pp

 Recessive Epistasis
Epistasis due to recessive genes is called recessive epistasis

 White coat produced by a recessive gene aa Gene B dominant state produces grey coat colour called agouti, and when recessive (bb) leads to black coat colour. The recessive gene for albinism (aa) is found to be epistatic to the gene for agouti (BB and Bb), and also to its recessive, homozygous allele (bb) for black.

 Duplicate gene When two or more genes have the same effect on a given trait, they are referred to as duplicate genes Yellow endosperm is dominant over white Two genes control the expression, yellow When both are in recessive condition it will produce white endosperm.

Cytoplasmic Inheritance
Uniparental inheritance: progeny inherit organelle genes exclusively from one parent. Maternal inheritance. Reason? Heteroplasmons : Normal as well as mutated organelle is present in the same zygote Cytoplasmic segregation : segregation of wild and mutated organelle to different daughter cells are called as cytoplamic segregation.

Linkage

Variation
Discontinuous Continuous

Nilsson-Ehle Experiment
deep red white

Deep Red 1

dark red 6

red 15

medium red 20

light red 15

very light red 6

white 1

Variation
Discontinuous
Individuals can be grouped in distinct class Controlled by one or few genes Not affected by environmental factors
Eg. Blood groups

Continuous
Difficult to group in distinct group Controlled by many genes Mostly affected by environment When plotted as a histogram, these data show a typical bellshaped normal distribution curve, with the mean, mode and median all being the same.
Eg. Height, colour,

Nilsson-Ehle's experiment on wheat kernel color

Changes in genetic material

Variation Chromosomal aberrations Mutation

Structural

Numerical

Numerical Aberration
Euploidy : Addition or removal of a complete set of chromosome Aneuploidy : Addition or removal of individual chromosome

 Euploidy: In euploidy an organism acquires an additional set of chromosomes over and above the diploid complement Triploid (3n), tetraploid (4n); pentaploid (5n), and hexaploid (6n)
Autopolyploids : If source of extra chromosome are same species Allopolyploids : If source of extra chromosome are Different species

 Autotriploid : Mostly sterile Eg. seedless watermelons, European pears and apples. Once formed, the triploid plants are healthy and robust and are propagated through asexual cuttings

 Autotetraploids; either fertile or only partially sterile Eg. Groundnut, potatoes and coffee

 Allopolyploidy Eg. The hybrid produced between radish and cabbage

hexaploid Triticum (2n + 44) X diploid Secale (2n = 14) to produce : Octaploid Triticale

 Polyploidy in agriculture:
Wheat : Einkorn, Emmer and Vulgare Einkorn (diploid 2n = 14). Not used for consumption Emmer (Tetraploid). Hybrid of Einkorn and Goat grass.
T. durum, T. dicoccum

 Vulgare (Hexaploid) T. aestivum (bread wheat) (AABBDD; 2n = 42) Originated through hybridisation between T. dicoccum (AABB) and a different species of goat grass followed by chromosome doubling

Aneuploidy
2n+1 : Trisomy 2n+2 : Tetrasomy 2n- 1 : Monosomy 2n-2 : Nullisomy

 Klinefelter syndrome XXY (Sterile male with low IQ), XYY Normal male Triplo-X trisomics (XXX) are phenotypically normal and fertile females Down syndrome 21st trisomy Edward syndrome 18th trisomy Pataus syndrome 13th trisomy Turner Syndrome XO (sterile female)

 Cause of Aneuploidy?
Mitotic or meiotic Non-disjunction

Structural aberrations
DELETION : Deletion of a small portion of a chromosome resulting in loss of one or more genes. It originates from breakage occurring at random in both chromatids of a chromosome (called chromosome break), or only in one chromatid (chromatid break).
Terminal : Single break at the end of the chromosome Interstitial : Two breaks occur in a middle portion of the chromosome. Multigenic deletion : several genes are mission due to deletion. More lethal than Interstitial

 Restitution: If the two raw ends of the same chromosome unite and restore the structure Exchange union: if two single breaks occur in two different chromosomes in a cell, the deleted segment of one chromosome may unite with the raw broken end on the other chromosome; this is called exchange union

 Fate of the deleted segment:

 Deletion loop: Pseudodominance :

 Eg: Cri du chat

heterozygous deletion of 5p15.2 and 5p15.3, of chromosome 5.

Williams syndrome.
An autosomal dominant syndrome characterized by unusual development of the nervous system and certain external features. Frequency 1 in 10,000 people. Patients often have pronounced musical or singing ability. Caused by a 1.5 Mb deletion on one homolog of chromosome 7.

Duplications
A chromosome duplication is a mutation in which part of the chromosome has been doubled
Tandem Reverse tandom Displacement duplication
Displaced on same arm Displaced on different arm

Origin: Unequal crossing over

 Eg: Bar eye mutant in Drosophila

The Bar mutant results from a small duplication on the X chromosome, which is inherited as an incompletely dominant, X-linked trait
779

358

68

45

Inversion
Inversions result when there are two breaks in a chromosome and the detached segment becomes reinserted in the reversed order
Paracentric: both breaks occur in one arm of the chromosome it leads to a paracentric inversion Pericentric : when a break occurs in each of the two arms, the centromere is included in the detached segment and leads to a pericentric inversion

Translocations
A translocation entails the movement of genetic material between nonhomologous chromosomes Nonreciprocal translocations: genetic material moves from one chromosome to another without any reciprocal exchange Reciprocal translocation : A two-way exchange of segments between the chromosomes

 Effect of Phenotype: May create new linkage relations that affect gene expression (a position effect) Chromosomal break can disrupt gene function
Eg. Neurofibromatosis

 Robertsonian translocation

Mutation
A mutation is defined as an inherited change in genetic information
Mutation is the source of all genetic variation Mutation is the cause of evolution Mutation helps organisms to survive in a particular conditions Mutation can be the reason for disease and disorder Mutations serve as important tools of genetic analysis Mutations are also useful for probing fundamental biological processes

Types of Mutations
Somatic mutations Germ-line mutations

 Mutation can be spontaneous or induced Point mutations

Alterations of single base pairs of DNA or of a small number of adjacent base

Types:
Base substitutions : Alternation of a single nucleotide in the DNA Insertions and deletions (Indel): The addition or the removal, respectively, of one or more nucleotide pairs

 Base substitution: Transition or Transversion

 Indel Mutations:

 Expanding trinucleotide repeats:

Fragile X syndrome; the most common hereditary cause of mental retardation The FMR 1 gene mutation) has 60 or fewer copies of this trinucleotide CGG, but in persons with fragile-X syndrome, the allele may harbor hundreds or even thousands of copies

 Forward mutation: A mutation that alters the wild-type phenotype, Reverse mutation : A reversion that changes a mutant phenotype back into the wild type.

 Suppressor mutations:
Is a genetic change that hides or suppresses the effect of an other mutation A suppressor mutation occurs at a site that is distinct from the site of the original mutation; thus, an individual organism with a suppressor mutation is a double mutant, possessing both the original mutation and the suppressor mutation but exhibiting the phenotype of an unmutated wild type

Intragenic suppression Mutation: happens on the same gene but different nucleotide or codon
Intergenic suppression Mutation: Happens on different gene

 Lethal Mutation: mutation that causes death of the organism

Recessive lethal Dominant lethal Semi lethal
Hemophilia

http://www.nature.com/scitable/topicpage /mendelian-ratios-and-lethal-genes-557

 Mechanism of point mutation: Base replacement

Tautomerization due to ionization Normal keto form change to imino or enol form

 Base analogues
The mutagen 5-bromouracil (5-BU) is an analogue of thymine and binds to adenine under normal conditions 5-BU frequently change to either the enol form or an ionized form that can bind to guanine

 Mechanism of point mutation: Base alteration Some mutagens causes specific mispairing by adding alkylating agents ;
eg Ethylmethanesulfonate (EMS) Adds ethyl group Nitrosoguanidine (NG) adds methyl group

Transposable Elements
Transposable elements are mobile DNA sequences that insert into many locations within a genome and often cause mutations and DNA rearrangements.
They make up at least 50% of human DNA

Transposition is the movement of a transposable element from one location to another. Short, flanking direct repeats of 3 to 12 base pairs are present on both sides of most transposable elements

Structure
In bacteria:
Insertion sequences (IS): Simple transposable elements Size 700 to 2000bp Contains one or two genes that code for transposase Flanking direct repeats

 Trasposones (Tn):
Larger than IS May carry genes other than transposase between the inverted sequences Some are with Tra gene helps in transfer to other bacteria during conjugation (conjugative transposons)

Integrones:
Integrones are transposons that can capture and express genes from other sources They are highly selective in insertion It contains , integrase, transposase, a gene cassette

Eukaryotic TE
Ty elements in yeast
These elements are retrotransposons

Ac element in maize
Ac elements are about 4500 bp long, including terminal inverted repeats of 11 bp, and the flanking direct repeats that they generate are 8 bp in length Each Ac element contains a single gene that encodes a transposase enzyme

 Ds element in maize
Ds elements are Ac elements with one or more deletions that have inactivated the transposase gene Ds elements can transpose in the presence of Ac elements

Mechanism of trasposition
Conservative:

Replicative:

 Transposition through an RNA intermediate (Retrotransposition)

SEX DETERMINATION
The mechanism by which sex is established is termed sex determination
Chromosomal Sex-Determining Systems Environmental Sex Determination Other mechanisms

Chromosomal Sex-Determining Systems

XX/XY sex-determination system

Most familiar, as it is found in humans. Females have two of the same kind of sex chromosome (XX, Homogametic) Males have two distinct sex chromosomes (XY, heterogametic). Y chromosome carries factors responsible for triggering male development

 XX-XO sex determination

In the grasshoppers In this system, females have two X chromosomes (XX), Males possess a single X chromosome (XO). O means no chromosome

 ZZ-ZW sex determination

W chromosome carries factors responsible for female development Male are homogametic (ZZ) Female are heterogametic (ZW) Eg Birds

Environmental Sex Determination

Marine mollusk Crepidula fornicata (slipper snail )

 This type of sexual development is called sequential hermaphroditism; each individual animal can be both male and female, although not at the same time. In Crepidula fornicata, sex is determined environmentally by the limpets position in the stack.

Red-eared slider turtle

American alligator

Alligator snapping turtle

Other mechanisms
Other insects, including honey bees and ants, use a haplodiploid sex-determination system. In this case diploid individuals are generally female, and haploid individuals (which develop from unfertilized eggs) are male. This sex-determination system results in highly biased sex ratios, as the sex of offspring is determined by fertilization rather than the assortment of chromosomes during meiosis

Sex determination in Drosophila

Total 8 chromosomes
3 pairs of autosomes 1 pair of sex chromosome 6A XX in Female and 6A XY in Male.

Here presence of the Y chromosome does not determine maleness in Drosophila; Flys sex is determined by a balance between genes on the autosomes and genes on the X chromosome. This type of sex determination is called the genic balance system.

 Here number of genes seem to influence sexual development. The X chromosome contains genes with female producing effects, whereas the autosomes contain genes with male-producing effects. Consequently, a flys sex is determined by the X:A ratio, the number of X chromosomes divided by the number of haploid sets of autosomal chromosomes. An X:A ratio of 1.0 produces a female fly; an X:A ratio of 0.5 produces a male.

Dosage Compensation
In XX/XY system, female has two X chromosome and male has one X. As a result X will over express causes problem in development. In animals there are different mechanism to reduce the effect of extra X chromosome, and are called as dosage compensation. By dosage compensation animals equalizes the amount of protein produced by X-linked genes in the two sexes.

Dosage compensation in Humans

Lyon hypothesis (Mary Lyon, 1961): within each female cell, one of the two X chromosomes randomly becomes inactive; If a cell contains more than two X chromosomes, all but one of them is inactivated by methylation. X inactive-specific transcript or XIST gene is active in future inactivated X (Xi).