遗传技术

Recombinant 重组 DNA is DNA that has been made by joining together DNA from two different sources. Genetic engineering 基因工程 is the deliberate changing of an organism's genetic material — often by transferring a gene 基因 into an organism so that the gene is expressed (switched on to make its protein 蛋白质).

The gene to be transferred can be obtained in three ways:

• cut out of the DNA of a donor 供体 organism,
• made from the donor's mRNA, using the enzyme reverse transcriptase 逆转录酶,
• built chemically from nucleotides 核苷酸.

The tools

A promoter 启动子 often has to be transferred along with the gene. The promoter is the "switch" that lets the gene be transcribed in its new organism, so without it the gene would stay silent.

To check the gene has gone in and is working, scientists add a marker gene 标记基因 next to it — for example one that codes for a fluorescent 荧光 (glowing) product. If the cells glow, the transfer worked.

Gene editing

Gene editing 基因编辑 is a precise form of genetic engineering. It inserts, deletes or replaces DNA at an exact site in the genome 基因组.

Copying and sorting DNA

• the polymerase chain reaction 聚合酶链式反应 (PCR) is used to clone 克隆 and amplify 扩增 DNA — to make millions of copies. It repeats cycles of heating and cooling, using a heat-stable enzyme called Taq polymerase.

• gel electrophoresis 凝胶电泳 separates DNA fragments 片段 by length. The fragments move through a gel in an electric field, and shorter fragments move further, so the lengths spread out into bands.

• microarrays 微阵列 are used to study whole genomes and to detect which genes are switched on, by picking up their mRNA.
• databases 数据库 store the nucleotide sequences of genes and the amino acid 氨基酸 sequences of proteins, so scientists anywhere can compare them.

Recombinant human proteins

A human gene can be put into bacteria or other cells so that they make a recombinant human protein — an exact copy of the human one. This is safer and never in short supply, and it avoids using proteins taken from animals or donors. Examples are insulin 胰岛素 (for diabetes 糖尿病), factor VIII (for haemophilia) and adenosine deaminase (for a faulty immune system).

Genetic screening

Genetic screening 基因筛查 tests a person's DNA for disease alleles before symptoms appear. Examples are the BRCA1 and BRCA2 alleles (which raise the risk of breast cancer 乳腺癌), Huntington's disease 亨廷顿病, and cystic fibrosis 囊性纤维化. Knowing the result helps people make informed choices about treatment and family.

Gene therapy

Gene therapy 基因治疗 treats a genetic disease by putting a working copy of a gene into the patient's cells. It has been used for SCID (a disease in which the immune system fails) and for some inherited eye diseases.

Social and ethical questions

Genetic screening and gene therapy raise concerns: who should see your genetic results, whether insurers or employers could misuse them, whether changes are safe and permanent, and who decides. These ethical 伦理 and social questions must be weighed against the benefits.

Genetic engineering can help feed a growing world by improving farmed animals and crops. Examples of genetically modified organisms 转基因生物 (GMOs) are:

• GM salmon 鲑鱼 that grow to size faster.
• soybean 大豆 made resistant to a herbicide 除草剂, so weeds can be sprayed without harming the crop.
• cotton 棉花 made resistant to insect pests 害虫, because it makes a protein that kills the insects.

GMOs also raise ethical and social questions: whether they are safe to eat, what effect they have on the environment and wild species, whether the engineered genes might spread, and whether a few large companies should control the food supply.

Worked example. A tiny DNA sample from a crime scene must be amplified and then compared with a suspect's. Outline the two techniques and what each achieves. PCR copies the DNA in repeated cycles: denaturation at about $95\ °\text{C}$ separates the strands, annealing at about $55\ °\text{C}$ lets primers 引物 bind at each end of the target, and extension at about $72\ °\text{C}$ has Taq polymerase build the new strands. Each cycle doubles the amount, so $n$ cycles give $2^n$ copies - 30 cycles turn one molecule into roughly a billion. Gel electrophoresis then separates the fragments: DNA is negatively charged because of its phosphate groups, so it moves towards the anode, and shorter fragments travel further through the gel. Matching band patterns point to the same source. Taq polymerase is used because it is thermostable - an ordinary polymerase would denature at $95\ °\text{C}$ in the very first cycle.

Common mistakes.

• Cutting the gene and the plasmid with different restriction enzymes. Use the same restriction enzyme, so both have complementary sticky ends that can pair before ligase seals them.
• Saying DNA ligase cuts DNA, or that restriction enzymes join it. Restriction enzymes cut; ligase joins the sugar–phosphate backbones.
• Saying larger fragments move further in gel electrophoresis. Shorter fragments move further, because they pass through the gel more easily.
• Saying DNA moves to the negative electrode. DNA is negatively charged (phosphate groups), so it moves towards the positive electrode (anode).
• Leaving the primers out of a PCR outline. Taq polymerase can only add nucleotides to an existing strand, so a primer must bind at each end of the target first.
• Forgetting the promoter. A gene moved into a new organism is not transcribed unless a suitable promoter is moved with it.
• Saying gene therapy changes the children of the patient. Gene therapy on body cells affects only the patient; the faulty allele can still be passed on in the gametes.

• Outline the toolkit: restriction enzymes (cut at specific sequences, sticky ends), ligase (join), plasmid vectors, and PCR (amplify).
• Explain gel electrophoresis: DNA separates by size, smaller fragments travel further — used in genetic fingerprinting.
• Give a balanced benefit vs concern for GMOs and gene therapy.