Biomed III Semester 2 exam

Genotype

The genetic code of a person. Ex. DNA Makeup

Phenotype

The way someone physically looks Ex. Tall/Short

Different types of genetic disorders

• Recessive

• Dominant

• Sex-Linked

• Multifactorial

• Chromosomal

• MItochondrial

Recessive

Both parents are carriers-they each have one recessive copy of a gene

Dominant

Both parents are dominant for that specific gene which guarantees that the child will receive it

Sex-Linked

Located on X chromosome and mothers are carriers (XX)

Nondisjunction

a failure of homologous chromosomes or sister chromatids to separate properly during cell division

Multifactorial

Caused by a combination of environmental factors and mutations in multiple genes

Multiactorial Gene Examples

• Alzheimers

• Heart Disease

• Breast Cancer

Chromosomal

Problems with missing or extra copies of genes along breaks/deletion or rejoining of chromosomes

What is Trisonomy 21

Down Syndrome

Chromosomal Disease Examples

Trisonomy 21

Mitochondrial

Caused by mutations in non chromosomal DNA found in the mitochondria. This is unique as it is solely passed from mother to child.

Mitochrondrial Disorder Example

Leber Hereditary Optic Neruopathy

Karyotype

The number and visual appearance of chromosomes in the cell nuclei of an organism or species

Homozygous Dominant

Same allele from each parent-Dominant trait is passed

Homozygous Dominant Example

TT

Homozygous Recessive

Same allele from each parent-Recessive trait is passed

Homozygous Recessive Example

tt

Heterozygous

There is one recessive and one dominant trait.

Heterozygous Example

Tt

PCR

Polymerase Chain Reaction

What does PCR do?

Creates a large # of copies of one gene for genetic testing.

Primers

Identifies the start and end if target sequence (gene)

Taq Polymerase

Builds DNA compliment of target sequence using dNTP’s.

dNTP’s

Free DNA nucleotides(base-A,T, G,C)

Stages of PCR

1. Denaturation

2. Annealing

3. Extension

Denaturation

94*C-Seperates DNA Strands

Annealing

40-65*C-Primer attaches

Extension

72*C-the dNTP’s are added and extends the DNA strand

What equation do you use to calculate the number of copies made?

2ⁿ⁺¹

What does n stand for in the equation to calculat the # of copies made

n represents the # of cycles

Electrophoresis

a laboratory technique used to separate DNA, RNA or protein molecules based on their size and electrical charge

How does Electrophoresis work?

Electrophoresis separates molecules, like DNA or proteins, based on their charge and size by using an electric field to move them through a gel or other medium.

How do you interpret the results of Electrophoresis?

To interpret electrophoresis results, observe band position, intensity, and the presence of expected bands

SNPs

Single Nucleotide Polymorphisms

What are SNP’s?

parts of the human genome that differ based on one nucleotide that make us unique

How to analyze sequence differences?

use sequence alignment tools like BLAST or aligners like LASTZ to compare sequences and identify regions of similarity and dissimilarity.

How do you use Electrophoresis to determine genotype and phenotype?

can be used to determine both genotype and phenotype by separating DNA fragments based on size, allowing scientists to analyze genetic variations and their corresponding traits. 

How do you select the correct restriction enzyme?

• the desired size and orientation of the resulting DNA fragments, the compatibility of the enzyme's cutting site with the desired location in the plasmid, and the presence of other restriction sites within the gene of interest or vector

• There’s rules

Types of Genetic Testing

• Carrier Screening

• Prenatal

• Newborn

• Ultrasound

• Amniocentesis

• Chorionic Villi Sampling

Carrier Screening

Test for one or both parents when you are planning pregnancy or are pregnant. Tested from blood/saliva and also tissue from the cheek

Prenatal

Fetus is tested for genetic disorders. Occurs 1st and 2nd trimester. Combination of fetal ultrasound and maternal blood test.

Newborn

Newborn is tested when it is 1 to 2 days old

Ultrasound

a medical imaging technique that uses high-frequency sound waves to create images of internal organs and tissues

Amniocentesis

Sample token from amniotic fluid to see if baby has any confitions. 15

Chrorionic Villi Sampling

Sample taken from placenta cells to test for baby conditions-10—13 weeks

Types of Diagnostic Imaging

• X-Ray

• CT Scan

• MRI

• Bone Scan

• PET Scan

X-Ray

2D images, dense structures, electromagnetic radiation

CT/CAT Scan

cross sectional images, bones and soft tissue, ionizing radiation

MRI (Magnetic Resonance Imaging)

cross sectional images, detailed soft/hard tissue, magnets

Bone Scan

2D images, dense structures-bones, tracers are used

PET (Positron Emission Technology) Scan

soft tissue-heart/organs, tracers, often combined with MRI or CT

Cancer

disease in which cells grow and divide uncontrollably, destroying healthy tissue

Biopsy

an examination of tissue removed from a living body to discover the presence, cause, or extent of a disease.

Cell Cycle

It involves growth, DNA replication, and the subsequent partitioning of cellular components into two daughter cells through a process called cell division.

Malignant

Cancer that spreads and impairs organs

Benign

Cancer that stays in the same place

Apoptosis

Programmed cell death

Normal Tissue

The tissue has normal cellular arrangement and cell structure.

Abnormal Tissue

Hyperplasia. The tissue contains a higher than usual number of cells. However, cell structure and the orderly arrangement of cells are normal.

Abnormal Tissue

Mild dysplasia. The tissue shows loss of normal tissue arrangement and cell structure, which is not extreme, thus is not considered cancerous.

Abnormal Tissue

Carcinoma in situ/Severe dysplasia. The tissue shows uncontrolled growth of cells and abnormal tissue arrangement and cell structure. However, the abnormalities remain within the original location.

Cancer

The tissue shows uncontrolled growth of cells and abnormal tissue arrangement and cell structure, which do not remain within the original location.

Normal Cells

• Divide in an organized matter

• specialized cells contain distinct features that enable them to perform their specific function

• Have the same size and shape

• One small nucleus

• Arranged in an organized matter and have well defined tissue boundaries

Cancer Cells

• Proliferate indefinetly

• lose their specialized cell features, which prevents them from performing their functions properly.

• vary in size and shape

• Large, variably shaped nuclei

• a disorganized arrangement of cells and tissue boundaries are poorly defined.

Cancer genes

• Proto-oncogene

• Tumor Suppressor Gene

• p53 Gene

• Oncogene

Proto-oncogene

(positive regulators) may be over activated or always on

Tumor Supressor Genes

(negative regulators) also may be inactivated (can’t turn off). Prevent tumors from forming

p53 Gene

Activates DNA repair proteins, halts cell cycle, triggers apoptosis

Oncogene

Can divide without growth factor signals. Ras mutation in pancreatic cancer

What gene mutation produce an Oncogene?

Mutation of proto-oncogene

Steps to producing a Microarray

1. isolating RNA,

2. creating labeled cDNA,

3. hybridization,

4. washing,

5. and image analysis.

What does Green mean on microarray?

the gene is strongly repressed or expressed at a lower level in the experimental sample compared to the control sample

What does Red mean on micorarray?

a gene is strongly expressed or overexpressed in one sample compared to another.

What does yellow mean on micorarray?

In a microarray, a yellow spot typically signifies that a gene is expressed at similar levels in both the experimental and control samples

Purpose of controls for Microarray?

They help identify potential issues in the microarray process, such as hybridization problems, RNA degradation, or amplification errors.

How to read results of microarray?

comparing the patient's DNA to a reference DNA, looking for differences in signal intensity

Cell Wall

Gives the cell its shape and surrounds the cytoplasmic membrane, protecting it from the environment. It also helps anchor appendages (pili/flagell) and making sure it doesn’t burst.

Cell Membrane

A layer of phospholipids and proteins which enclose the interior of bacterium. Their function is to regulate the flow of materials in and out of the cell.

Capsule

Third protective covering made of polysaccharides. Keep the bacterium from drying out and protect it from phagocytosis by larger microorganisms.

Nucleoid Region (DNA)

Region of the cytoplasm where the chromosomal DNA is located. Strand of DNA are found here.

Plasmid

Carry genes for replicating their DNA, transferring themselves from one host to another and a variety of phenotypes.

Ribosome

Microscopic “factories” found in all cells. They translate the genetic code from the molecular language of nucleic acid to that of amino acids. Distributed through cytoplasm.

Pili

Small Hairlike projections emerging from the outside cell surface. Help assist the bacteria to attach to other cells and surfaces. Without Pili bacteria can’t infect.

Flagella

Tail looking structure that provides a means of locomotion for bacteria.

Why are plasmids and bacteria used for genetic engineering?

• Carry genes for replicating

• Replicates independently

• Easily manipulated

• reproduction-Conjugtion

• Bacterial cells replicate fast

What is the role of Ca²⁺in transformation?

They interact with negative charges and create an electrostatically neutral situation.

What is the role of Ice incubation in transformation?

The cold temperature slows down cell membrane activity, allowing for better DNA binding, and prepares the cells for the subsequent heat shock.

What is the role of heat shock during transformation?

This creates a temperature imbalance on either side of the bacterial membrane, and sets up a current? With the “ionic shield” in place, then DNA can be swept through the adhesion zone.

What is the role of a restriction enzyme in transformation?

to cut both the donor DNA (containing the gene of interest) and the plasmid vector (often used to carry the donor DNA into a recipient cell) at specific, recognizable sequences

How do you determine if transformation is successful?

observing phenotypic changes, performing DNA analysis, conducting enzyme assays, using selection methods, and analyzing colonies. Specifically, this involves looking for visible differences in the organism's appearance

pGLO plasmid components

• Ori

• Bla

• Ara C

• GFP

Ori gene

Origin of replication

Bla Gene (beta-lactamase gene)

Ampicillin resistance (makes sure bacteria will grow ampicillin)

Ara C gene (Arabinose-C)

Regulates GFP transcription (makes sure GFP is produced)

GFP gene (Green Fluroscent Protein Gene)

“protein of interest” it makes sure the bacteria will glow under UV light

Will it grow or glow?-LB

Grow but No glow

Will it grow or glow?-LB/amp

Growth but No glow

Will it grow or glow?-LB/amp/ara

Growth and glow