A restriction enzyme recognizes GAATTC. Person X is heterozygous: one chromosome has this site intact between two flanking cuts (producing a 4 kb fragment), while the other chromosome has a mutation that destroys this internal site. What banding pattern appears when Person X's DNA is run on a gel?
ATwo bands: 4 kb and 7 kb
BOne band: 7 kb (the mutation always dominates on a gel)
CThree bands: one at 4 kb, one at 3 kb, and one at 7 kb
DNo bands — heterozygosity prevents clear fragment separation
A heterozygote has two alleles: the intact chromosome produces the 4 kb fragment (the enzyme cuts at both flanking sites and the internal site); the mutant chromosome lacks the internal site, so the enzyme produces only one larger fragment spanning the distance covered by the 4 kb + 3 kb segments — typically 7 kb. Both alleles' DNA is present in the sample, so the gel shows all products: the 4 kb and 3 kb bands from the intact chromosome AND the 7 kb band from the mutant chromosome — three bands total. Option A (just two bands) would describe a homozygous normal individual; only heterozygotes show three.
Question 2 Multiple Choice
Why do VNTR (minisatellite) loci produce individually unique banding patterns suitable for forensic identification, while a typical restriction site polymorphism at a single locus usually has only two alleles?
AVNTR loci are located in coding regions where mutations accumulate faster than at non-coding restriction sites
BVNTRs have a short tandem repeat motif that varies in copy number between individuals due to unequal crossing over and replication slippage, creating dozens to hundreds of distinct alleles rather than just two
CVNTR loci are near centromeres and therefore experience more mutation per generation
DRestriction site polymorphisms occur only once per genome, but VNTRs occur in multiple locations simultaneously
The power of VNTR-based fingerprinting comes from the mechanism of variation. Minisatellites consist of a short repeat unit (e.g., a 16 bp motif) that can occur in tandem 10, 20, 50, or 200+ times — and unequal crossing over or polymerase slippage during replication can increase or decrease this count. The result is dozens to hundreds of distinct alleles per locus. At multiple independent VNTR loci, the probability of two unrelated individuals sharing the same multi-locus pattern becomes astronomically small. A single-nucleotide restriction site polymorphism, by contrast, is biallelic — the site either exists or it doesn't — providing much less discriminatory power.
Question 3 True / False
RFLP analysis requires sequencing the individual's DNA to identify genetic differences, since the underlying single nucleotide changes are too small to detect by gel electrophoresis alone.
TTrue
FFalse
Answer: False
This is precisely what makes RFLP analysis powerful — it reveals genetic differences without sequencing. A single nucleotide change that creates or destroys a restriction site changes the size of the resulting fragment, and fragment size differences of even a few hundred base pairs are readily resolved by gel electrophoresis. The gel image translates molecular genetic variation directly into visible banding patterns. RFLP was developed specifically to detect polymorphisms efficiently before sequencing was fast or cheap, and the underlying principle remains: you're measuring the consequence of the mutation (fragment size) rather than reading the mutation itself.
Question 4 True / False
A single nucleotide change in a DNA sequence can produce a detectably different banding pattern on an agarose gel following restriction enzyme digestion.
TTrue
FFalse
Answer: True
If the single nucleotide change occurs within a restriction enzyme's recognition sequence, it eliminates (or creates) a cut site. This converts two smaller fragments into one larger fragment (or vice versa) — a size difference easily detected by gel electrophoresis. For example, changing GAATTC (EcoRI site) to GAACTC by a single transversion eliminates the cut site entirely, merging two bands into one larger band. This is the molecular basis of RFLP analysis: restriction fragment length polymorphisms exist because individuals differ in which restriction sites they carry, and those differences in restriction sites are often caused by single nucleotide changes.
Question 5 Short Answer
Explain how RFLP analysis could be used to identify the biological parents of an individual, even without sequencing their DNA.
Think about your answer, then reveal below.
Model answer: RFLP-based parentage testing works because restriction fragment patterns are heritable — a child must receive one allele at each locus from each parent. If you digest the child's DNA and both parents' DNA with the same restriction enzyme panel and run the fragments on a gel, each child's band must be present in at least one parent at every polymorphic locus. A 7 kb band in the child that appears in the mother's profile but not the father's confirms maternal inheritance; a 4 kb band in the child that matches the alleged father but no other candidate provides paternity evidence. The more polymorphic loci analyzed, the lower the probability that a non-parent could match by chance. For minisatellite VNTR-based fingerprinting, multi-locus profiles can establish parentage with virtual certainty.
Parentage testing illustrates Mendelian logic applied at the molecular level. Each RFLP band corresponds to an allele inherited from one parent or the other. Exclusion (a band in the child that cannot be in either parent) rules out parentage; inclusion across many loci establishes it statistically. Modern STR-based testing uses the same logic but with PCR-amplified microsatellites, giving greater sensitivity and the ability to work with degraded or tiny samples — but the conceptual foundation is identical to Jeffreys' original RFLP approach.