amazon.co.ukA-Level Biology revision on gene expression and technology: transcription factors, epigenetics, stem cells, recombinant DNA, PCR, gel electrophoresis and genome projects, with multi-board exam practice and mark-scheme logic.
Every cell contains the same genome, yet only some genes are translated - control of gene expression explains differentiation and cell specialisation. Transcription factors are proteins that bind specific DNA sequences (promoters) and control whether RNA polymerase can transcribe a gene: stimulatory factors enable transcription; inhibitory factors block it. Oestrogen is the stock example of an external signal controlling transcription: it is lipid-soluble, diffuses through the cell-surface membrane, binds a complementary receptor (ER) in the cytoplasm, causing the receptor to change shape and release an inhibitory molecule; the oestrogen-receptor complex then enters the nucleus and binds promoter regions, acting as a transcription factor that activates target genes (e.g. in some breast cancers, over-expression drives uncontrolled cell division). siRNA controls expression after transcription: double-stranded siRNA unwinds and associates with RISC (RNA-induced silencing complex); one strand guides RISC to a complementary mRNA, which RISC cleaves, so translation cannot occur - RNA interference, usable therapeutically to silence disease genes.
Epigenetics is heritable change in gene expression without altering the base sequence, controlled by the epigenome - a layer of chemical tags on DNA and histones. Two mechanisms are examinable: DNA methylation - methyl groups (-CH₃) added to cytosine in CpG islands attract proteins that condense chromatin (heterochromatin), blocking RNA polymerase access and silencing transcription; histone acetylation - acetyl groups added to histone lysine residues loosen the DNA-histone packing (euchromatin), giving RNA polymerase access and activating transcription. Deacetylation condenses chromatin again. Epigenetic changes are heritable through cell division and sometimes between generations, and link environmental factors to disease: e.g. methylation changes in cancer (tumour-suppressor genes silenced, oncogenes activated), smoking-induced epigenetic marks. The epigenome is reversible - drugs can inhibit methylation enzymes or histone deacetylases, an A* discussion point. When asked to explain epigenetic control, always state the mechanism (methyl/acetyl), the chromatin state (condensed/loosened), and the outcome (transcription on/off) - three linked marks.
Epigenetic tags can be inherited across generations - examples studied include famine exposure (Dutch Hunger Winter) leaving methylation marks in offspring, and maternal care altering methylation of stress-response genes - showing how the environment in one generation changes gene expression in the next without changing the base sequence; this bridges genetics and evolution questions, since epigenetic variation is another heritable layer on which selection can act.
Stem cells are undifferentiated cells that divide by mitosis and differentiate. Classify them precisely - the three-tier definition earns full marks: totipotent - can differentiate into all body cell types plus extra-embryonic tissue (placenta); only the zygote and first few divisions (up to the 8-cell stage). Pluripotent - from the inner cell mass of the blastocyst; differentiate into all somatic cell types but not placental tissue; source of embryonic stem cells. Multipotent - e.g. bone marrow (haematopoietic) stem cells; differentiate into a limited range of related cells (blood cells). During differentiation, most genes are switched off (heterochromatin) so only genes needed for the cell's function are expressed - the mechanism linking stem cells to transcription control above. iPS cells (induced pluripotent stem cells) are adult somatic cells reprogrammed back to pluripotency by introducing extra transcription factors: they divide endlessly like embryonic stem cells but avoid destroying embryos, sidestepping the main ethical objection, and can be made from the patient's own cells to avoid immune rejection. Uses: bone marrow transplants for leukaemia/lymphoma, regenerative medicine (growing tissues/organs), studying differentiation, testing drugs without animal/human trials. Ethics debate expects balance: embryo destruction vs. medical benefit; consent and ownership of embryos; safety (uncontrolled division, tumour risk); alternatives (iPS, adult stem cells).
Expression is also controlled post-transcriptionally and post-translationally: splicing and RNAi (above), and protein activation (e.g. phosphorylation, as in the cAMP second-messenger cascade in homeostasis). In prokaryotes, the lac operon (OCR 6.1.1; WJEC) is the classic model: lactose (allolactose) binds the repressor, releasing it from the operator so RNA polymerase transcribes the structural genes (β-galactosidase, lactose permease); glucose scarcity raises cAMP which helps CAP binding - transcription only when lactose present and glucose absent. Learn the operon sequence if your board lists it (OCR and WJEC do; AQA does not).
Recombinant DNA combines DNA from different organisms; because the genetic code is universal, a gene transplanted between species still produces the same protein. Obtaining the target gene: (1) reverse transcriptase - isolate mRNA for the target protein (abundant in specialised cells, e.g. β-cells for insulin), use reverse transcriptase (from retroviruses) to make cDNA; cDNA contains no introns, so bacteria (which cannot splice) can translate it, and single-stranded cDNA is made double-stranded with DNA polymerase; (2) restriction endonucleases - cut the gene from DNA at a specific palindromic recognition site, leaving staggered sticky ends; (3) a gene machine synthesises the gene directly from a known sequence (from genome databases). Inserting into a vector: the same restriction enzyme cuts a plasmid at a single site, producing complementary sticky ends; gene and plasmid anneal by complementary base pairing, and DNA ligase seals the sugar-phosphate backbones with phosphodiester bonds, producing recombinant DNA. Transforming cells: plasmids are returned to host bacteria (e.g. E. coli) via Ca²⁺ and heat shock (or electroporation), making membranes permeable; marker genes identify successful uptake - fluorescent markers (GFP), antibiotic-resistance markers (survivors grown on antibiotic agar carry the plasmid) or enzyme markers; replica plating distinguishes plasmids with the target gene. Fermenters culture transformed bacteria to mass-produce protein (insulin, factor VIII, vaccines).
In vivo (vectors, bacteria) vs in vitro (PCR) comparison is a standard six-marker: in vivo is slow but accurate (proof-reading enzymes, low error rate), copies large fragments, produces modified proteins (post-translational modification possible in bacteria is limited - note insulin is made as pro-insulin), and carries little risk of contamination; in vitro (PCR) is rapid, automatable, needs only tiny DNA samples, but has a higher error rate (Taq lacks proof-reading) and produces many identical copies without modification. Ethics of recombinant DNA and transgenic organisms: benefits (mass cheap insulin, GM crops with higher yield/nutrition, gene therapy); concerns (antibiotic-resistance marker genes spreading to pathogens, GM gene flow to wild populations, animal welfare, corporate control of seeds); evaluation questions want both sides plus a justified conclusion.
PCR (polymerase chain reaction) amplifies DNA in vitro in three repeating stages: (1) denaturation at ~95 °C - hydrogen bonds break, separating the double helix into single strands; (2) annealing at ~55 °C - short single-stranded primers bind complementarily to the 3' ends of the target sequences, providing the starting point for DNA synthesis (and preventing re-annealing of the template strands); (3) extension at ~72 °C - Taq polymerase (thermostable, from the bacterium Thermus aquaticus) adds free nucleotides (dNTPs) to synthesise complementary strands. Each cycle doubles the DNA (2ⁿ after n cycles: ~1 billion copies after 30 cycles). A thermal cycler automates the temperature cycling. Exam questions ask for the role of primers and Taq exactly as above, and why Taq is needed (does not denature at 95 °C).
Gel electrophoresis separates DNA fragments by size: DNA fragments are negatively charged by their phosphate groups, so in an agarose gel with an electric current they migrate towards the anode (+); shorter fragments travel further/faster through the gel pores, separating fragments by length; fragments are visualised under UV with a fluorescent dye. Combined with PCR, electrophoresis underlies DNA profiling (genetic fingerprinting): amplify variable number tandem repeat (VNTR) regions, electrophorese, and compare banding patterns between individuals - used in forensics, paternity testing and diagnosis. DNA probes: short single-stranded DNA fragments labelled with radioactive or fluorescent tags; they hybridise to complementary target sequences on a membrane or gel, locating specific alleles - the basis of screening for mutated alleles (e.g. BRCA1) in medical genetics. DNA sequencing: chain-termination (Sanger) or modern high-throughput methods determine the exact base order of fragments; allele sequences from genome projects let probes be designed. AQA and OCR may also expect allele frequency screening and personalised medicine as applications.
Genome projects sequence the entire DNA of organisms (Human Genome Project completed 2003). Proteomics - the study of the complete set of proteins an organism produces - reveals that the ~20,000 human genes produce far more proteins via alternative splicing and post-translational modification, so knowing the genome does not fully predict the proteome. Sequencing pathogen genomes enables rapid vaccine/drug design and epidemiology; comparative genomics maps evolutionary relationships and identifies disease genes. Medical applications of sequencing and probes: screening for mutated alleles before symptoms (predictive testing), personalised medicine (drug choice and dosage by genotype), prenatal and preimplantation diagnosis, and locating genes for gene therapy.
Ethical evaluation is a guaranteed 5-6 marker on AQA (3.8.4) and Edexcel (T8) - structure it as benefits, concerns, conclusion. Benefits: earlier diagnosis and treatment; personalised medicine; gene therapy for inherited diseases (CF, SCID); GM crops addressing food security; forensic accuracy. Concerns: genetic discrimination (insurers/employers using screening data); privacy of genetic databases; consent and ownership; embryo use in therapy and screening (destruction of embryos, designer babies); safety of gene therapy (insertion mutagenesis, immune reactions); GM organisms escaping into ecosystems. Gene therapy: supplementing or silencing faulty genes - somatic therapy (body cells only, not inherited, e.g. CF inhalation therapy) vs germ-line therapy (germ cells, heritable, currently illegal in humans on ethical grounds). Screening ethics questions expect you to discuss autonomy, confidentiality, and the right not to know, with a justified judgement - quote the spec point that decision-making is an individual and societal matter.