5.1 Basic and Molecular Genetics

5.1 Basic and Molecular Genetics

Quick Answer: A locus is a chromosomal address; alleles are the sequences that sit there; antithetical antigens are the mutually exclusive protein products of those alleles (K/k, C/c, Fya/Fyb). Silent (amorphic) alleles produce no antigen (group O, deleted RHD). Cis means same haplotype; trans means the opposite chromosome — C in trans to RHD can weaken D. ISBT numbers antigens as a six-digit code; traditional names remain the bench language. Molecular methods (PCR-SSP, real-time PCR, sequencing, microarray) rescue typing when serology fails. They do not replace serology when a silencing mutation or RHD-CE-D hybrid sits outside the typed SNPs. Exam-classic chromosomes: ABO 9, RH 1, KEL 7, FY 1.

Blood-group genetics on the BB exam is not a biochemistry elective. It is why two people type K+k+ and a third types K−k+, why a genotype can say D+ while the tube says D−, and why you reach for a molecular assay after massive transfusion instead of believing the last gel card. Outline II.A (June 9, 2026) tests the vocabulary, the naming systems, the methods, and the limits.

Genes, alleles, loci, and antithetical antigens

A gene is the DNA sequence that encodes a blood-group protein or a glycosyltransferase. The locus is where that gene lives. An allele is one version of that gene. KEL01 and KEL02 are alleles at the KEL locus; they produce K and k. You inherit one haplotype from each parent. The phenotype is what serology sees. The genotype is the allele pair you carry.

Do not call K and k “genes.” They are antithetical antigens — products of alleles at the same locus. One haplotype cannot carry both K and k. A person who types K+k+ is a heterozygote (KEL01/KEL02). Classic antithetical pairs: K/k, Kpa/Kpb, Jsa/Jsb, C/c, E/e, Fya/Fyb, Jka/Jkb, M/N, S/s, Lua/Lub, Dia/Dib, Coa/Cob. A “K+k−” cell is almost always KEL01/KEL01, not a missing locus.

A silent or amorphic allele produces no antigen. Group O is the everyday ABO amorph: ABOO encodes a truncated, inactive transferase. Deleted RHD is the common European D− genotype. FY silencing alleles and KELnull (K0) are amorphs in their systems. Silent is not weak. Weak alleles (weak D, Kmod, Fy^x) still make some antigen; amorphs make none.

Cis versus trans is haplotype geometry. Two alleles are cis when they ride the same chromosome. They are trans when they sit on opposite homologs. Three exam-classic payoffs:

• Cis-AB: one chromosome encodes a transferase with both A and B activity. The person can type AB with a group O parent.
• C in trans to RHD (Ceppellini effect): a haplotype such as dCe (r') facing Dce (R0) or DcE (R2) can weaken D. The cell is still D+, but a weak-D card may look equivocal. C that is cis to D (R1 = DCe) does not produce that same weakening.
• Compound antigens such as Ce and f (ce) form when C and e, or c and e, are cis. A DCE/dce person has C and e in trans and is f-negative. Phenotype without haplotype thinking mis-predicts f, Ce, and G.

ISBT numeric names versus the names on your rack

The International Society of Blood Transfusion (ISBT) assigns each system a three-digit number and each antigen a three-digit number. Concatenated, they are a six-digit identifier. RH is system 004. D is antigen 001 of that system, so D is 004001 or RH1. C is RH2, E is RH3, c is RH4, e is RH5. KEL is 006: K is KEL1, k is KEL2. FY is 008 (FY1 = Fya). JK is 009. Traditional names — D, K, Fya, Jka — remain the language of panels, computer crossmatches, and BB stems. ISBT numbers appear when a stem wants you to decode a rare antigen or read a molecular report. You do not memorize the catalog. You do need to know that the first three digits are the system and that traditional and ISBT names are the same antigen, not two antigens.

Exam-classic chromosome locations — only these four are worth a flashcard:

Do not invent chromosomes for Kidd, MNS, or Lewis. The exam rewards these four addresses.

Molecular methods you must assign

PCR-SSP (sequence-specific primer PCR) uses a primer whose 3' end matches one allele. Amplification means the allele is present; no band means it is not. It is fast and excellent for common SNPs (K/k, Fya/Fyb, Jka/Jkb, S/s). It interrogates only the nucleotides you aimed at.

Real-time PCR adds a fluorescent probe and reads amplification as it happens. Copy-number assays estimate RHD zygosity (one versus two RHD genes) — the number a prenatal counselor needs when the father of a D− mother’s fetus is D+. Real-time assays on maternal plasma detect fetal RHD (cell-free fetal DNA) and can spare RhIG when the fetus is D−.

Sequencing (Sanger for a targeted exon, next-generation sequencing for the gene) reads the actual sequence. It classifies a novel RH variant, distinguishes weak D type 1 from a partial D, and finds a mutation a SNP chip never printed. Order it when phenotype and SNP genotype disagree.

Microarray / bead-chip HEA platforms interrogate dozens of blood-group SNPs in one well. They are the workhorse for extended antigen prediction in chronically transfused patients, especially sickle-cell programs, and for RH variant prediction in African-ancestry patients whose serologic D, C, and e look ordinary but whose RHD and RHCE are hybrid.

When molecular typing is the right next test

Serology is first-line for ABO/Rh and antibody identification. Molecular typing earns its keep in five BB-classic settings:

1. Weak D and partial D. Serologic weak D can be a quantitative weak D (types 1, 2, 3 — common in European ancestry; most facilities treat as D+ for transfusion and as D+ for RhIG) or a partial D that lacks epitopes and can make anti-D. Only genotyping (or a validated partial-D serologic panel) separates them. Do not call every weak D “give D− blood and RhIG” if the laboratory’s policy uses genotype to reclassify types 1, 2, and 3.

2. Prenatal typing. Fetal RHD from maternal plasma, paternal RHD zygosity, and fetal KEL1 when the mother has anti-K. You cannot type fetal red cells from a maternal venipuncture by serology.

3. Recently transfused patients. Circulating donor red cells make the phenotype a mixture. DNA from white cells or a buccal swab reports the patient’s alleles.

4. DAT-positive red cells. Bound IgG blocks typing reagents, especially at antiglobulin phase. Genotype bypasses the coat.

5. RH variant prediction. African-ancestry patients with sickle cell disease are enriched for RHD and RHCE hybrids. A serologic e+ may still make alloanti-e if that e is partial e.

Limitations — genotype does not always win

Molecular prediction fails in predictable ways. Silencing mutations outside the assayed SNP (splice-site, promoter, distant stop) leave an “antigen-positive” genotype and an antigen-negative cell. The GATA-1 promoter mutation that silences FYB on red cells is the teaching example — a chip that only reads the 125G>A Fya/Fyb SNP will call the person Fyb+ when the red cells are Fy(b−). Hybrid genes, especially RHD-CE-D hybrids, shuffle exons between RHD and RHCE. An assay aimed at exon 4 or 7 can call RHD present in a person who is serologically D− or partial D. RHDpsi (the 37-bp insertion pseudogene common in African D− donors) is the other classic false-positive RHD PCR.

Molecular typing is not a substitute for serology in all cases. ABO must still be determined serologically before transfusion. A molecular type does not replace an antiglobulin crossmatch when an antibody is present. New alleles, rare hybrids, and assay design limits mean you confirm unexpected results with serology, family study, or sequencing. If a stem gives you a microarray Fyb+ result and Fy(a−b−) red cells in an African-ancestry patient, the answer is the GATA-1 promoter silencing mutation, not “the chip is broken” and not “the patient is K0.”