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🎓 FULL LECTURE — Biochemistry & Genetics (Pages 51-59)

Think of it like this: I'm your professor. Let's go line by line.

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📖 PAGE 51 — CRISPR/Cas9 & Blotting Procedures

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🔬 CRISPR/Cas9

"A genome editing tool derived from bacteria."

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"Consists of a guide RNA (gRNA), which is complementary to a target DNA sequence..."

• Guide RNA (gRNA): A small piece of RNA (like a messenger molecule) that acts as a GPS. You design it to match the exact piece of DNA you want to find. It "guides" the system to the right location.
• Complementary: DNA is a double strand. Each strand pairs with the other - A pairs with T, and G pairs with C. "Complementary" just means "it matches/pairs perfectly."

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"...and an endonuclease (Cas9), which makes a single- or double-strand break at the target site."

• Endonuclease: A protein (enzyme) that cuts DNA. Think of it as the scissors.
• Cas9: The specific scissors protein from bacteria. Once the gRNA GPS leads it to the target, Cas9 cuts the DNA.
• Single-strand break: Cuts only one of the two DNA strands (like cutting one side of a ladder)
• Double-strand break: Cuts BOTH strands completely — the DNA is now in two pieces (like cutting the entire ladder in half)

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"Applications include removing virulence factors from pathogens, replacing disease-causing alleles of genes with healthy variants (eg, sickle cell disease), and specifically targeting tumor cells."

• Virulence factors: The "weapons" a germ uses to make you sick. CRISPR can cut those weapons out.
• Alleles: Different versions of the same gene. Think of a gene like a word - "colour" vs "color" are two versions (alleles). In sickle cell disease, there's a bad allele; CRISPR can replace it with the good version.
• Tumor cells: Cancer cells. CRISPR can be directed to cut and destroy cancer cell DNA specifically.

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The diagram shows two outcomes after CRISPR cuts DNA:

• NHEJ (Non-Homologous End Joining): The cell tries to repair the cut but makes errors - like taping two pieces of paper together but crookedly.
• Frameshift: DNA is read like a sentence in groups of 3 letters (codons). If the repair adds or removes letters, the whole sentence is scrambled - the gene is broken.
• Knock-out: The gene is silenced/destroyed. Good for disabling a bad gene.

• HDR (Homology Directed Repair): More precise repair - the cell uses a "template" (donor DNA you provide) to repair the cut correctly.
• Knock-in: You're inserting a new, healthy gene. Good for replacing a bad gene with a good one.

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🧪 Blotting Procedures (SNoW DRoP - MEMORIZE THIS!)

• Southern = DNA
• Northern = RNA
• Western = Protein

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Southern Blot

"DNA sample is enzymatically cleaved into smaller pieces, which are separated by gel electrophoresis, and then transferred to a membrane."

"Membrane is exposed to labeled DNA probe that anneals to its complementary strand."

• DNA probe: A small, known piece of DNA that is "labeled" (made to glow or be detectable). You design it to be complementary to the sequence you're looking for.
• Anneals: Pairs up with / binds to. The probe hunts through your membrane and sticks only to its matching sequence.

"Resulting double-stranded, labeled piece of DNA is visualized when membrane is exposed to film or digital imager."

• Once the probe binds, you expose the membrane to film (like an X-ray) or scan it. Wherever the probe bound, a dark band appears. That band = your target DNA.

"Useful to identify size of specific sequences (eg, determination of heterozygosity, # of CGG repeats in FMR1 to diagnose Fragile X syndrome)"

• Heterozygosity: Whether you have two different versions of a gene (one from mom, one from dad). On the blot, you'd see two differently sized bands instead of one.
• Fragile X Syndrome: A genetic disorder where one gene (FMR1) has too many repeated copies of the letters "CGG." Southern blot can count how many repeats are there to diagnose this.

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Northern Blot

"Similar to Southern blot, except that an RNA sample is electrophoresed. Useful for studying mRNA levels and size, which are reflective of gene expression. Detects splicing errors."

• Same process as Southern blot, but instead of DNA, you're analyzing RNA (specifically mRNA - the "messenger" that carries instructions from DNA to make proteins).
• Why is this useful? Because the amount of mRNA = how active a gene is. If a gene is "turned on," it makes lots of mRNA.
• Splicing errors: After mRNA is made, it gets processed (edited). Northern blot can detect if this editing went wrong (which can cause disease).

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Western Blot

"Sample protein is separated via gel electrophoresis and transferred to a membrane. Labeled antibody is used to bind relevant protein. This helps identify specific protein and determines quantity."

• Same idea, but for proteins.
• Instead of a DNA probe, you use an antibody (a detection molecule that binds only to the specific protein you're looking for).
• Uses: Confirm HIV diagnosis, identify specific proteins in disease research.

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📖 PAGE 52 — Flow Cytometry, Microarrays, ELISA, Molecular Cloning

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🔬 Flow Cytometry

"Laboratory technique to assess size, granularity, and protein expression (immunophenotype) of individual cells in a sample."

• Immunophenotype: The "identity card" of a cell - what proteins are on its surface. Different cell types wear different surface proteins, like uniforms.
• Granularity: How grainy/complex the inside of a cell is. Immune cells that contain granules (like neutrophils) are more granular.

"Cells are tagged with antibodies specific to surface or intracellular proteins. Antibodies are then tagged with a unique fluorescent dye."

• You take your blood/tissue sample.
• Add antibodies that stick ONLY to specific proteins on certain cell types.
• Those antibodies are tagged with a fluorescent dye - a glowing color.
• Different antibodies = different colors. So different cell types glow different colors.

"Sample is analyzed one cell at a time by focusing a laser on the cell..."

• Each cell is passed single-file through a laser beam (like a checkout scanner at a supermarket).
• When the laser hits a glowing cell, a detector reads: how big is it? How granular? What color (which proteins)?

"Data are plotted either as histogram (one measure) or scatter plot (any two measures)"

• Histogram: A bar graph. Shows how many cells have a certain level of one marker.
• Scatter plot: A dot plot showing two markers at once. Each dot = one cell.

The CD3/CD8 example in the diagram:

• CD3: A marker found on ALL T-cells (a type of immune cell)
• CD8: A marker found only on killer T-cells (cytotoxic T cells)
• Left lower quadrant (CD3-, CD8-): Not T-cells at all
• Right upper quadrant (CD3+, CD8+): Killer T-cells (they have both markers)
• Left upper quadrant (CD3+, CD8-): Helper T-cells (have CD3 but not CD8)

"Commonly used in workup of hematologic abnormalities (eg, leukemia, paroxysmal nocturnal hemoglobinuria, fetal RBCs in pregnant person's blood) and immunodeficiencies (eg, CD4+ cell count in HIV)."

• Leukemia: Cancer of blood cells - flow cytometry identifies abnormal cells
• Paroxysmal nocturnal hemoglobinuria (PNH): A disease where red blood cells lack protective proteins - flow cytometry detects this
• CD4+ cell count in HIV: The virus destroys CD4+ helper T-cells; flow cytometry counts how many are left to monitor disease severity

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🔬 Microarrays

"Used to compare the relative transcription of genes in two RNA samples. Can detect single nucleotide polymorphisms (SNPs) and copy number variants (CNVs) for genotyping, clinical genetic testing, forensic analysis, and genetic linkage analysis when DNA is used."

• Transcription: The process of making RNA from DNA. "Relative transcription" = comparing how active genes are in two different samples (eg, cancer cell vs. normal cell).
• Single nucleotide polymorphisms (SNPs): Tiny variations in DNA - one letter is different from person to person (eg, position 1000 is A in most people but G in some). These variations can increase disease risk.
• Copy number variants (CNVs): Some people have 2 copies of a gene, others have 1 or 3. This variation can cause disease.
• Think of microarrays like: A giant checklist (chip) with thousands of DNA spots. You pour your RNA/DNA sample on it; it glows wherever there's a match, revealing which genes are active or what variations exist.

• Nocturnal hemoglobinuria, fetal hemoglobin, leukemia: Hematology
• RBCs in pregnant person's blood: Detecting fetal red blood cells in mom's circulation
• HIV: Monitoring the virus

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🔬 Enzyme-Linked Immunosorbent Assay (ELISA)

"Immunologic test used to detect the presence of either a specific antigen (in direct ELISA) or antibody (in indirect ELISA) in a patient's blood sample."

• Antigen: A foreign substance (like a virus protein) that triggers an immune response.
• Antibody: A protein your immune system makes to fight a specific antigen.
• Direct ELISA: Looking for an antigen in the blood (eg, is the HIV protein here?)
• Indirect ELISA: Looking for antibodies (eg, has the body made anti-HIV antibodies? = the person was infected)

"Detection involves the use of an antibody linked to an enzyme, producing a detectable signal. Can have high sensitivity but lower specificity than Western blot. Often used to screen for HIV infection."

• Enzyme-linked: The detection antibody has an enzyme attached. When it finds its target and binds, the enzyme reacts with a substance to produce a visible color change (like a pregnancy test turning pink).
• Sensitivity vs. Specificity:
  • High sensitivity = catches almost all true positives (good screening test - don't miss cases)
  • Lower specificity = may have more false positives (needs confirmation with Western blot)
• That's why: First you screen with ELISA, then confirm with Western blot.

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🧬 Molecular Cloning

"Production of a recombinant DNA molecule in a bacterial or eukaryotic cell line host. Useful for production of human proteins in bacteria (eg, human growth hormone, insulin)."

• Recombinant DNA: DNA made by combining DNA from two different sources (eg, human gene inserted into bacteria's DNA).
• The idea: Take the gene for human insulin → insert it into bacteria → bacteria reads the gene and makes human insulin → you harvest it.
• Before recombinant DNA technology, diabetic patients used insulin from pigs and cows. Now we make identical human insulin using bacteria. Cheaper, safer, unlimited supply.
• Eukaryotic cell line: Sometimes bacteria can't make complex human proteins properly, so you use more complex cells (like yeast or mammalian cells) as the "factory."

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📖 PAGE 53 — Karyotyping & Fluorescence In Situ Hybridization (FISH)

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🔬 Karyotyping

"Colchicine is added to cultured cells to disrupt the assembly of mitotic spindles and arrest cells at mitosis."

• Mitosis: Cell division. During cell division, chromosomes condense and become visible.
• Mitotic spindles: Protein cables that pull chromosomes apart during cell division.
• Colchicine: A drug (originally from crocus flowers) that destroys these spindle cables, freezing the cell mid-division. This keeps chromosomes visible and accessible.
• Why? Chromosomes are only visible (as distinct X-shaped structures) during cell division. Colchicine keeps them "frozen" in that visible state.

"Chromosomes are stained, ordered, and numbered according to morphology, size, arm length ratio, and banding pattern (arrows in normal cell, cancer cell)."

• Chromosomes are stained with dyes → they develop unique banding patterns (like barcodes).
• These bands help identify each chromosome uniquely.
• The 23 pairs are then arranged in order from largest to smallest — this arranged picture is the karyotype.

"Can be performed on a sample of blood, bone marrow, amniotic fluid, or placental tissue."

• Amniotic fluid: Fluid surrounding the baby in the womb. Contains fetal cells. Used for prenatal diagnosis of chromosomal abnormalities.
• Bone marrow: Used to diagnose blood cancers (leukemia).

"Used to diagnose chromosomal imbalances (eg, autosomal trisomies, sex chromosome abnormalities)."

• Trisomy: Having 3 copies of a chromosome instead of 2. Example: Down syndrome = Trisomy 21 (3 copies of chromosome 21).
• Sex chromosome abnormalities: eg, Turner syndrome (45,X - missing one X), Klinefelter syndrome (47,XXY).

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🔬 Fluorescence In Situ Hybridization (FISH)

"Fluorescent DNA or RNA probe binds to specific gene or other site of interest on chromosomes. Used for specific localization of chromosomal anomalies."

• FISH = a more precise version of karyotyping. Instead of looking at all chromosomes broadly, you design a glowing probe to find ONE specific gene or chromosome region.
• The probe glows (fluoresces) wherever it binds → you can see exactly where that gene is, or if it's missing/duplicated.

"Microdeletion - no fluorescence on a chromosome compared to fluorescence on the second copy of that chromosome corresponds to the fluorescence signal that corresponds to tetrasomy (chromosome 5)."

• Microdeletion: A tiny piece of a chromosome is missing - too small to see on regular karyotype, but FISH catches it.
• Translocation: A piece of one chromosome moved to another chromosome.
• Duplication: An extra copy of a chromosomal segment.

• Velocardiofacial syndrome (chromosome 22 microdeletion)
• DiGeorge syndrome
• Specific translocations in leukemia

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📖 PAGE 54-55 — Gene Expression, RNA Interference, Genetic Terms

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🧬 Gene Expression Modifications (Transgenic Mice)

"Knock-out = removing a gene, taking it out. Knock-in = inserting a gene."

• Knock-out mouse: Delete a gene → see what happens without it → understand its function
• Knock-in mouse: Insert a human disease gene → mouse develops human disease → test drugs
• Random insertion: Gene is inserted randomly. It's always "on" (constitutive expression).
• Targeted insertion: Gene is inserted at a specific location. Can be turned on/off conditionally.

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🧬 RNA Interference

"Process whereby small non-coding RNA molecules target mRNAs to inhibit gene expression."

"Naturally produced by cell as hairpin structures. Loose nucleotide pairing allows broad targeting of related mRNAs. When miRNA binds to mRNA, it blocks translation of mRNA and sometimes facilitates its degradation."

• Hairpin structure: The miRNA folds back on itself into a hairpin shape.
• Loose pairing: miRNA doesn't have to match perfectly → it can silence multiple related genes at once (broad targeting).
• Blocks translation: mRNA carries the instructions to make protein. If miRNA blocks mRNA, no protein is made.
• Abnormal miRNA in cancer: Tumors can overactivate certain miRNAs that silence tumor suppressor genes (the body's brakes on cancer). No brakes = uncontrolled growth.

"Usually derived from exogenous dsRNA source (eg, virus). Once inside a cell, siRNA requires complete nucleotide pairing, leading to highly specific mRNA targeting. Results in mRNA cleavage prior to translation."

• Exogenous: From outside the cell (eg, a virus injects double-stranded RNA).
• Complete pairing: siRNA is like a precision sniper - matches only ONE specific mRNA.
• mRNA cleavage: The mRNA is physically cut and destroyed = gene is silenced.
• Medical use: siRNA drugs can silence specific disease-causing genes (eg, patisiran for hereditary amyloidosis).

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📚 Genetic Terms (Definitions & Examples)

Codominance

"Both alleles contribute to the phenotype of the heterozygote."

• Phenotype: What you can observe/measure (eg, blood type, height, eye color).
• Heterozygote: Has two different alleles (one from each parent).
• In codominance, BOTH alleles are expressed simultaneously.
• Example: Blood groups A, B, AB, α-antitrypsin.
  • If you get allele A from one parent and allele B from the other → you get blood type AB (both are expressed, neither dominates).

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Variable Expressivity

"Not all individuals with a pathogenic gene variant show the disease variant to the same degree. % penetrance × probability of inheriting variant."

• Pathogenic gene variant: A gene mutation that can cause disease.
• Same mutation can cause severe disease in one person and mild symptoms in another.
• Example: Neurofibromatosis type 1 (NF1) - some people have just a few café-au-lait spots, others have hundreds of tumors.
• Example: Two patients with neurofibromatosis type 1 may have varying disease severity.

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Incomplete Penetrance

"% penetrance = probability of inheriting variant. % penetrance × probability of expressing the phenotype."

• Penetrance: Whether you show the disease at all.
• If penetrance is 80%, 20% of people who carry the mutation will never show symptoms, even though they carry it.
• Example: BRCA1 gene mutations do not always result in breast or ovarian cancer. Not every BRCA1 carrier gets cancer (but their risk is much higher).

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Pleiotropy

"One gene contributes to multiple phenotypic effects."

• One mutation → affects multiple organs/systems.
• Example: Cystic fibrosis manifests with thick mucus in the lungs and GI tract, pancreatic insufficiency, and infertility.
  • One broken gene (CFTR) → lungs get clogged, pancreas fails, sweat is salty, male infertility. One gene, many effects.

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Anticipation

"Increased severity or earlier onset of disease in succeeding generations."

• Each generation, the disease gets worse or starts earlier.
• Why? Some genes have repeat sequences (like a stutter - CAGCAGCAG...) that expand (get longer) each generation. More repeats = worse disease.
• Example: Trinucleotide repeat diseases (eg, Huntington's disease, myotonic dystrophy). If grandpa got Huntington's at 60, dad may get it at 45, and son may get it at 30.

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Loss of Heterozygosity

"If a patient inherits or develops a mutation in a tumor suppressor gene, the wild-type allele must be deleted/mutated before cancer develops. This is not true for oncogenes."

• Tumor suppressor gene: The body's brakes on cancer. You need BOTH copies to work.
• Wild-type: The normal, working version.
• Think of it like two brake lines in a car. You need to cut BOTH for the brakes to fail.
• If you inherit one broken copy (first hit) and then the second copy gets damaged later (second hit) → both brakes fail → cancer. This is the "two-hit hypothesis."
• Example: Retinoblastoma (eye cancer in children) - the RB gene. First hit inherited, second hit acquired → cancer.
• Oncogenes: Accelerators for cell growth. You only need ONE copy to be broken = disease. (Not the two-hit rule.)

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Epistasis

"The allele of one gene affects the phenotypic expression of alleles in another gene."

• One gene "overrides" or modifies another gene.
• Example: Albinism. Even if you have genes for dark hair/skin color, if the gene that MAKES pigment is broken → you produce no pigment at all. The pigment production gene is "epistatic" to the color genes.
• Aneuploidy: Abnormal number of chromosomes due to chromosomal nondysjunction during mitosis (eg, Down syndrome - trisomy 21, Turner syndrome, oncogenesis).

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More Genetic Terms (Page 55):

Dominant Negative Mutation

"Exerts a dominant effect. A heterozygote produces a nonfunctional altered protein that also prevents the normal gene product from functioning."

• Normally, if one copy of a gene is broken, the other copy compensates.
• In dominant negative, the broken copy makes a BAD protein that also BLOCKS the good protein.
• Like a broken key that not only doesn't open the door but also gets stuck in the lock preventing the good key from working.
• Example: p53 tumor suppressor gene mutation. The mutant p53 protein blocks normal p53 from binding DNA and doing its job.

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Linkage Disequilibrium

"Tendency for certain alleles to occur in close proximity on the same chromosome more or less expected by chance."

• Genes that are physically close on a chromosome tend to be inherited together (they rarely get separated during reproduction).
• Example: HLA gene, CFTR gene - certain combinations of variants at nearby genes tend to travel together in populations.

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Mosaicism

"Presence of genetically distinct cell lines in the same individual. Somatic mosaicism - mutation only in certain tissues."

• Not every cell in your body has the same DNA mutation.
• Early in development, one cell mutated. All cells descended from that one cell have the mutation. Other cells don't.
• Like a quilt: Most patches are one pattern, but some are different.
• Germline mosaicism: The mutation is in the egg or sperm cells → can be passed to children even if the parent appears unaffected.
• Example: McCune-Albright syndrome - due to G-protein activating mutation. Presents with unilateral café-au-lait spots, polyostotic fibrous dysplasia, and precocious puberty. Lethal if the mutation occurs before fertilization (affecting all cells), but survivable in mosaicism (affecting at least one endochondropathy).

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Locus Heterogeneity

"Mutations at different loci result in the same disease."

• Same disease, different genes causing it.
• Example: β-thalassemia, albinism, familial hypercholesterolemia. Different mutations in different genes can all cause the same clinical picture.

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Allelic Heterogeneity

"Different mutations in the same locus result in the same disease."

• Multiple different mutations WITHIN the same gene can cause the same disease.
• Example: mDNA passed from mother to all children - hereditary optic neuropathy.

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Uniparental Disomy

"1 parent and no copies from the other parent. Offspring receives 2 copies of a chromosome from one parent."

• Instead of getting one chromosome 15 from mom and one from dad, you get both from the SAME parent.
• Why it matters: Some genes are imprinted - they're only expressed if they come from a specific parent. If both copies come from the wrong parent, those imprinted genes won't work.
• Heterodisomy: Both copies from same parent are different (one of each of that parent's chromosomes).
• Isodisomy: Both copies are identical (same chromosome duplicated).

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Heteroplasmy

"Presence of both normal and mutated mtDNA, resulting in variable expression in mitochondrially inherited disease."

• Mitochondria have their own DNA (separate from nuclear DNA).
• If some mitochondria have the mutation and others don't, the severity of disease depends on what proportion are mutated.
• Different tissues in your body may have different proportions = variable symptoms.

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📖 PAGE 56 — Hardy-Weinberg Principle & Imprinting Disorders

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📐 Hardy-Weinberg Principle

"In a given population where mating is at random, allele and genotype frequencies will be constant from generation to generation."

• p + q = 1 (p = frequency of dominant allele A, q = frequency of recessive allele a)
• p² + 2pq + q² = 1 (frequencies of genotypes AA, Aa, aa)

• p² = proportion of people who are homozygous dominant (AA)
• 2pq = proportion of heterozygous carriers (Aa)
• q² = proportion of homozygous recessive (aa) = people who show the recessive disease

• No mutation
• Completely random mating
• No net migration
• No natural selection
• Large population

• q² = 1/3200 → q = 1/56.6 ≈ 0.017
• p = 1 - 0.017 ≈ 0.983
• Carrier frequency (2pq) = 2 × 0.983 × 0.017 ≈ 1/29 (about 3.5% of population are carriers)

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🧬 Disorders of Imprinting

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Prader-Willi Syndrome

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Angelman Syndrome

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📖 PAGE 57 — Modes of Inheritance

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🧬 Autosomal Dominant

"Often due to defects in structural genes. Many generations, both males and females are affected."

• Only one bad copy of the gene is needed to cause disease.
• Passes through EVERY generation (doesn't skip).
• Males and females equally affected.
• Every affected person has at least one affected parent.
• Risk to children of affected person: 50% chance each child is affected.

• Often pleiotrophic (one gene, many effects)
• Variable expressivity - severity differs between people
• Family history crucial
• With 1 affected parent: each child has 25% chance AA, 50% Aa (affected), 25% aa (unaffected)

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🧬 Autosomal Recessive

"Often due to enzyme deficiencies. Usually seen in only 1 generation."

• Both copies of the gene must be broken to cause disease.
• Carriers (one bad copy) are healthy.
• Disease often appears in ONE generation (skips generations because carriers don't show symptoms).
• Parents of an affected child are usually carriers (Aa × Aa).
• Each child of two carriers: 25% AA (normal), 50% Aa (carrier), 25% aa (affected disease).
• Increased risk in consanguineous families (relatives marrying relatives, because rare alleles concentrate).

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🧬 X-Linked Recessive

"Commonly more severe in males. Females usually must be homozygous to be affected."

• The gene is on the X chromosome.
• Males only have ONE X chromosome (XY) → if that X has the bad gene, they have the disease.
• Females have TWO X chromosomes (XX) → they need BOTH copies bad to show disease (usually just carriers).
• No male-to-male transmission (a father gives his Y chromosome to sons, not his X).

• Carrier mother (X^N X^a) × normal father (X^N Y):
  • Sons: 50% affected (X^a Y), 50% normal (X^N Y)
  • Daughters: 50% carriers (X^N X^a), 50% normal (X^N X^N)

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🧬 X-Linked Dominant

"Transmitted through both parents. Children of affected mothers have 50% chance of being affected. 100% of daughters and 0% of sons of affected fathers will be affected."

• The mutation on X chromosome causes disease even with ONE copy.
• Affected fathers pass it to ALL daughters (they give their X to daughters) but NONE of their sons (sons get Y from dad).
• Examples: Fragile X syndrome, hypophosphatemic rickets (also called X-linked hypophosphatemia)

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🧬 Mitochondrial Inheritance

"Transmitted only through the mother. All offspring of affected females may show signs of disease."

• Mitochondria are inherited entirely from the mother (sperm contributes no mitochondria).
• Affected MOTHER → can pass to ALL children (sons and daughters).
• Affected FATHER → passes to NO children.
• Variable expression due to heteroplasmy.

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📖 PAGE 58 — Autosomal Dominant Diseases, Autosomal Recessive Diseases, Cystic Fibrosis

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🦴 Autosomal Dominant Diseases (List)

• Achondroplasia - dwarfism due to FGFR3 gene mutation
• Familial hypercholesterolemia - high cholesterol from birth
• ADPKD (autosomal dominant polycystic kidney disease) - kidneys fill with cysts
• Hereditary hemorrhagic telangiectasia (Osler-Weber-Rendu) - abnormal blood vessels
• Huntington's disease - progressive brain degeneration
• Marfan syndrome - tall, long fingers, heart problems
• Neurofibromatosis type 1 and 2 - nerve tumors
• Tuberous sclerosis - benign tumors in multiple organs
• Von Hippel-Lindau disease - blood vessel tumors

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⚗️ Autosomal Recessive Diseases (List)

• Albinism, PKU, sickle cell, cystic fibrosis, Tay-Sachs, hemochromatosis, Wilson disease - all AR
• Mucopolysaccharidoses (except Hunter syndrome which is X-linked)
• Glycogen storage diseases, Kartagener syndrome
• ARPKD - autosomal RECESSIVE polycystic kidney disease

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💨 Cystic Fibrosis (MAJOR TOPIC - KNOW IN DETAIL)

Genetics

"Autosomal recessive, defect in CFTR gene on chromosome 7. Most common lethal genetic disease in patients with European ancestry."

• CFTR gene: Cystic Fibrosis Transmembrane Conductance Regulator
• Most common mutation: ΔF508 (deletion of phenylalanine at position 508)
• AR disease - you need TWO bad copies

Pathophysiology (How it causes disease)

"CFTR encodes an ATP-gated Cl- channel (secretes Cl- in lungs/GI tract, reabsorbs Cl- in sweat glands). Phe508 deletion → misfolded protein → improper protein trafficking → protein absent from cell membrane → Cl- and HCO3- secretion → compensatory Na+ reabsorption via epithelial Na+ channels (ENaC) → Na+ reabsorption → more negative transepithelial potential difference → Cl- concentration in pilocarpine-induced sweat test → alkalosis and hypokalemia."

1. CFTR is a chloride channel in the cell membrane - it lets Cl- ions move in and out.
2. In lungs: CFTR secretes Cl- out → water follows by osmosis → keeps mucus thin and watery.
3. In sweat glands: CFTR reabsorbs Cl- back into the body (so sweat isn't too salty).
4. When CFTR is broken: In lungs, Cl- can't get out → water doesn't follow → mucus becomes thick and sticky → bacteria can't be cleared → infections.
5. In sweat glands, Cl- can't be reabsorbed → sweat becomes very salty → this is the basis of the sweat chloride test (>60 mEq/L = diagnostic for CF).

Diagnosis

"Can present with contraction alkalosis and hypokalemia (ECF H₂O/Na⁺ loss via sweating and concomitant renal K/H⁺ wasting). ↑ immunoreactive trypsinogen (newborn screening); ↑ Cl- concentration in pilocarpine-induced sweat test."

• Sweat chloride test: Pilocarpine is applied to skin to stimulate sweating. CF patients have >60 mEq/L chloride in sweat (vs <40 in normal).
• Newborn screening: Blood test for immunoreactive trypsinogen (a pancreatic enzyme - elevated because pancreatic ducts clog up).

Complications

"Recurrent pulmonary infections (eg, S. aureus infancy and early childhood, P. aeruginosa adulthood), allergic bronchopulmonary aspergillosis (ABPA), chronic bronchitis and bronchiectasis, reticulonodular pattern on CXR, opacification of sinuses, nasal polyps, nail clubbing. Pancreatic insufficiency, malabsorption with steatorrhea, and fat-soluble vitamin deficiency (A, D, E, K). Meconium ileus in newborns."

• P. aeruginosa: A specific bacteria that colonizes CF lungs - particularly dangerous, hard to treat.
• Bronchiectasis: Permanent widening of airways due to recurrent infections and inflammation.
• Pancreatic insufficiency: Thick mucus clogs pancreatic ducts → digestive enzymes can't reach the gut → fat malabsorption → foul-smelling, fatty stools (steatorrhea) → deficiency of fat-soluble vitamins A, D, E, K.
• Meconium ileus: Newborn CF babies can't pass their first stool because it's too thick - a pediatric emergency.
• Male infertility: Congenital absence of vas deferens (tubes carrying sperm).
• Females may be subfertile due to thick cervical mucus.

Treatment

"Multidirectional: chest physiotherapy, aerosolized dornase alfa (DNase), and inhaled hypertonic saline. Ibuprofen for anti-inflammatory effect. Pancreatic enzyme replacement therapy. CFTR modulators (potentiators or correctors). Azithromycin prevents acute exacerbations."

• DNase: Breaks down DNA in mucus (dead white blood cells release DNA, making mucus even thicker).
• Hypertonic saline: Draws water into the airway → thins mucus.
• CFTR modulators: NEW drugs that actually fix the broken CFTR protein:
  • Potentiators (eg, Ivacaftor): Keep the CFTR channel open longer (for gating mutations).
  • Correctors (eg, Lumacaftor, Tezacaftor): Help the misfolded protein reach the cell surface (for ΔF508).
  • Elexacaftor/Tezacaftor/Ivacaftor (Trikafta): Triple combination - works for most CF patients.

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📖 PAGE 59 — X-Linked Recessive Diseases, Muscular Dystrophies

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🧬 X-Linked Recessive Diseases (List)

• Bruton agammaglobulinemia (no B-cells, recurrent infections)
• Hemophilia A (Factor VIII deficiency) and B (Factor IX deficiency)
• G6PD deficiency
• Fabry disease (sphingolipid storage)
• Ornithine transcarbamylase (OTC) deficiency
• Wiskott-Aldrich syndrome
• Lesch-Nyhan syndrome
• Hunter syndrome (MPS type II)
• X-inactivation (Lyonization): In females, one X chromosome is randomly inactivated in each cell. If the activated X has a bad gene, that cell shows the disease effect. This is why female carriers can sometimes show mild symptoms (skewed inactivation).

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💪 Muscular Dystrophies — Duchenne & Becker

Duchenne Muscular Dystrophy (DMD)

"X-linked recessive disorder typically due to frameshift deletions or nonsense mutations → truncated or absent dystrophin protein → progressive myofiber damage."

• Dystrophin gene (DMD): The LARGEST protein-coding human gene.
• Frameshift mutation: A deletion or insertion that shifts the reading frame → completely broken protein.
• Dystrophin: A protein that anchors muscle fibers to the extracellular matrix (scaffolding). Without it, muscle fibers tear during contraction.
• Progressive myofiber damage: With every contraction, fibers tear. Over time, muscle is replaced by fat and connective tissue.

"Weakness begins in pelvic girdle muscles and progresses superiorly. Pseudohypertrophy of calf muscles due to fibrofatty replacement of muscle. Onset before 5 years of age. Dilated cardiomyopathy is common cause of death."

• Pseudohypertrophy: The calf muscles LOOK big and muscular, but they're actually full of fat and scar tissue - not real muscle. Like a balloon filled with garbage instead of air.
• Gowers' sign: Child uses hands to "walk up" their own legs to stand up from the floor (due to weak pelvic/hip muscles). Classic exam finding.
• Lordosis: Exaggerated inward curve of the lower back (compensating for weak hip muscles).
• Dilated cardiomyopathy: Heart muscle also lacks dystrophin → heart becomes weak and dilated.

• ↑ CK (Creatine Kinase): Released from damaged muscle cells - a blood marker.
• Aldolase: Another enzyme elevated from muscle damage.
• Genetic testing: Confirms diagnosis.

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Becker Muscular Dystrophy

"X-linked recessive disorder typically due to non-frameshift deletions in dystrophin gene (partially functional dystrophin - Becker is better). Less severe than Duchenne. Onset in adolescence or early adulthood."

• In Becker, the deletion is in-frame (doesn't shift the reading frame) → dystrophin is made, but it's shorter or slightly abnormal → still partly functional.
• Key difference from Duchenne: Becker patients can walk into adulthood; Duchenne patients lose walking by age 12.
• Same genes, different mutation types → very different severity. A perfect example of allelic heterogeneity.
• Deletions can cause both Duchenne and Becker, with large deletions spanning one or more exons.

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Myotonic Dystrophy

"Autosomal dominant. Onset age 20-30 years. CTG trinucleotide repeat expansion in the DMPK gene."

• Myotonia: Inability to RELAX muscles after contracting. Grip something → can't let go.
• CTG repeat expansion: The stuttering mutation that gets worse each generation (anticipation).

• Cataracts: Lens opacification in a relatively young person.
• Toupee sign: Early frontal balding in males.
• Gonadal atrophy: Testes shrink.
• Difficulty releasing hand from handshake (classic!)
• Muscle wasting, testicular atrophy, cardiac arrhythmia.
• Muscle biopsy shows ring fibers and central nuclei (normally nuclei are at the edge of muscle fibers).

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🎯 QUICK SUMMARY TABLE

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🩺 KEY DISEASE MEMORY AIDS

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