17 Genetics and Development 1 Flashcards by Echo Quinton

Describe general features of the human genome and the chromosomal basis of inheritance (mitosis and meiosis)

3.2 billion DNA base pairs
20,000 protein-encoding genes (1.5% of genome)
Mitosis: no change in ploidy (diploid parent produces diploid daughter)
Meiosis: Diploid parent produces four haploid gametes

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Interpret gene location using ISCN nomenclature (eg 6p21.1)

ISCN Nomenclature
- Numbered consecutively from centromere to telomere
- Ex 6p21.1
- Chromosome 6
- p = short arm
- Region 2
- Band 1
- Sub-band 1

ISCN = international system for Human Cytogenetic Nomenclature

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Describe basic principles of cytogenetics (karyotyping)

Karyotyping is the process by which a karyotype is prepared from photographs of chromosomes, in order to determine the chromosome complement of an individual, including the number of chromosomes and any abnormalities.
1. Sample collection and tissue culture.
2. Arresting cells at metaphase.
3. Swelling, separating and spreading chromosomes using hypotonic solution.
4. Separating chromosomes onto the slide.
5. Staining or banding.
6. Arranging the results- a karyotype.

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Define genetics

Study of genes and their roles in inheritance

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Define genomics

Study of all a persons genes, including interactions of those genes with the persons environment

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Define epigenetics

study of reversible changes in the chromatin landscape as determinants of gene function rather than on changes of the genomic sequence itself
ex DNA methylation, histone modifications

Epigenetic mechanisms DO NOT alter the underlyiing DNA sequence

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Define Ploidy
-Euploidy?
-Aneuplody?
-polyploid?

Ploidy: Number of chromosome sets in a cell
- Euploidy: normal (ie 46, XY/ 46XX)
- Aneuploidy: abnormal (ie 47, XY, +21) Male with 47 chromosomes (extra copy of 21)
- Polyploid: 3+ chromosome sets (ie triploid, tetraploid, pentaploid etc) // Not usually compatible with life (ft in cancer cells)

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Define mosaicism
How might it arise?

2 or more cell lines in an individual
Common cause is nondisjuction in an early postzygotic mitotic division

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Contrast genetic disease with congenital disease

Congenital Disease
-Present at birth
-Congenital = born with
-Does not imply or exclude a genetic basis for birth defect
-1/25 babies have congenital defects (canada)
Genetic Disorder = Specific Genetic Cause
-Single gene disorders with large effects
(eg sickle cell anemia, Cystic fibrosis)
-Chromosomal disorders
(eg Trisomy 21 // Chromosome 22q11.2 deletion (digeorge syndrome))
-Complex multigenetic disorders
(Diabetes, schizophrenia, crohn disease)

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Classify genetic disorders (3 types)
Examples of each

Single-gene disorders: where a mutation affects one gene. Sickle cell anemia is an example.
Chromosomal disorders: where chromosomes (or parts of chromosomes) are missing or changed. Chromosomes are the structures that hold our genes. Down syndrome, DiGeorge syndrome (22q11.2)
Complex (multigenetic) disorders: where there are mutations in two or more genes. Often your lifestyle and environment also play a role. Diabetes, schizophrenia, crohn disease Colon cancer is an example.

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What is a GENE?

Basic physical unit of inheritance
Sequence of nucleotides in DNA that encodes either RNA or protein

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During which phase of cell division are chromosomes visible?

Metaphase chromosomes are visible

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Metaphase chromosomes are classified by (?)

Metaphase chromosomes are classified by the position of the centromere:
- Telocentric
- Acrocentric
- Submetacentric
- Metacentric

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What is g-banding?

G- banding (Giemsa banding)
-technique used in cytogenetics to produce a visible karyotype by staining condensed chromosomes

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Metaphase chromosomes are classified by the position of the centromere:
What are the four types of chromosome?

Telocentric
Acrocentric
Submetacentric
Metacentric

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Submetacentric chromosome

-centromere present near the middle and divides the chromosome into two unequal arms.
- It is an L-shaped chromosome.
Most chromosomes in humans are submetacentric, e.g. 2nd, 4th to 12th, 17th, 18th, and X chromosomes.

Telocentric chromosome

-centromeres present at the end of the chromosome.
-It is not observed in humans.
-The shorter p arm is barely visible.
-The shape of the chromosome at the anaphase is like the letter ‘i’.

Metacentric chromosome

-centromere separates the chromosome into two equally sized arms and is located in the center of each one.
-The metacentric chromosome is shaped like an X.
E.g. 1st, 3rd, 16th, 19th, and 20th are metacentric chromosomes in humans.

Acrocentric Chromosome?

centromere is present near the end of chromosomes.
It forms a very short p arm and a very long q arm.
E.g. 13th, 14th, 15th, 21st, 22nd, and Y chromosomes are acrocentric in humans.

Limitation of karyotyping? Other tests that can be done?

Cannot detect small rearrangements (<5Mb) Regions where banding is not distinctive
Other tests: FISH , Microarray

FISH = Fluorescence in situ hydridization

What is FISH?

Fluorescence in situ hydridization
- molecular cytogenetic technique that allows the localization of a specific DNA sequence or an entire chromosome in a cell.

What is Microarray?

A microarray is a laboratory tool used to detect the expression of thousands of genes at the same time.
- DNA microarrays are microscope slides that are printed with thousands of tiny spots in defined positions, with each spot containing a known DNA sequence or gene
- can identify small duplications or deletions of genetic material that previously could not be identified using conventional karyotyping alone
- important for Microdeletion

What is Sanger Sequencing? how does it differ from PCR?

Sanger sequencing, aka the “chain termination method”, is a method for determining the nucleotide sequence of DNA
- uses a DNA sequence of interest as a template for a PCR that adds modified nucleotides, called dideoxyribonucleotides (ddNTPs), to the DNA strand during the extension step
- Basic priciples of molecular assays
- Sanger sequencing differs from PCR in that only a single primer is used in the reaction.

What is real time PCR?

Real-time PCR is the technique of collecting data throughout the PCR process as it occurs, thus combining amplification and detection into a single step. This is achieved using a variety of different fluorescent chemistries that correlate PCR product concentration to fluorescence intensity (1)

Restriction fragment length analysis? What is an RFLP probe?

RFLP analysis: form of genetic testing to observe whether an individual carries a mutant gene for a disease that runs in his or her family - An **RFLP probe** is a *labeled DNA sequence* that *hybridizes* with one or more fragments of the digested DNA sample after they were separated by gel electrophoresis, thus revealing a *unique blotting pattern characteristic to a specific genotype at a specific locus*. - Short, single- or low-copy genomic DNA or cDNA clones are typically used as RFLP probes. | RFLP = Restriction Fragment Length Polymorphism ## Footnote uses specific restriction endonucleases

What is Single-base primer extension?

Single-base extension (SBE) is a method for determining the identity of a nucleotide base at a specific position along a nucleic acid - an effective and sensitive tool that can type over 30 known loci scattered throughout an organism's genome in a single reaction.

What is Next Generation Sequencing

Next-generation sequencing (NGS) is a massively parallel sequencing technology that offers ultra-high throughput, scalability, and speed. The technology is used to determine the order of nucleotides in entire genomes or targeted regions of DNA or RNA.

Next generation sequencing vs Sanger sequencing? - NGS, the genetic material (DNA or RNA) is **?**, to which **?** of known sequences are attached, through a step known as **?**, enabling the fragments to **?** The bases of each fragment are then identified by **?**. - *The main difference between* Sanger sequencing and 2G NGS stems from **?**, with **?** allowing the processing of millions of reactions in parallel, resulting in **?, ?, ?** and **?**

NGS, the genetic material (DNA or RNA) is fragmented, to which oligonucleotides of known sequences are attached, through a step known as **adapter ligation**, enabling the fragments to interact with the chosen sequencing system. The bases of each fragment are then identified by their emitted signals. *The main difference between* Sanger sequencing and 2G NGS stems from **sequencing volume**, with NGS allowing the processing of millions of reactions in parallel, resulting in high-throughput, higher sensitivity, speed and reduced cost

Amplicon?

an amplicon is a piece of DNA or RNA that is the source and/or product of amplification or replication events

What is Cri Du Chat Syndrome?

- Cat's cry syndrome - unusual high pitched cry - 5p deletion syndrome - Microcephaly, hypertelorism, epicathal folds, low set ears, sometimes with preauricular tags and micrognathia - Overal incidence estimated to be as high as 1/15000 births - 46,XX,del5p15