Shanghai dogs are very special!

1. Shanghai dogs are very special!

2. There are many dogs in Shanghai Xuhui JTU campus

4. Hypothesis： Shanghai dogs can read 汉字 (and obey orders)

5. Result： The clever dog 狗recognized the label 肉 and ignored the box labeled 菜 Conclusion: At JTU even the dogs are clever

6. What was good? Any problems? What should we learn?

7. What was good? Good observation Great idea!

8. Any problems? 鬣狗 hyenas are not dogs How can you find out whether the bacterium that is growing in your flask is the correct one? Discuss 老托的故事 I have seen many scientists wasting time because they failed to recognize contaminants.

9. How to detect contaminants? • Streak out for single colonies • (Phase contrast) microscopy • Test for known markers (e.g. antibiotic resistance) • Use specific antibodies etc. • PCR (only on pure cultures) • Check early and keep checking! PNAS2001vol. 98/14 The extent of cell line contamination is enormous. 45 out of 252 (1/5) new cell cultures deposited at a German cell line repository were contaminated by another cell line. Hundreds of scientific reports based on fraudulent [wrong] cell lines were published.

10. Any problems? Living things have several senses to ‘read’ the environment List all the senses you know, and the organisms where they are most important. Start with bacteria, give examples. Try to explain without using the dictionary

11. 5 4 3 If you repeat an experiment, make sure that it is a true, independent repeat 2 1 2 2 2 1 1 2

12. Does a specific mutation increase antibiotic production? Wild-type strain 0 Gene replacements to generate identical mutations Mutant strain 1 Clonal copy of 1 Mutant strain 2 Growing the bacteria (culture broth/medium, flask/fermenter) Measuring antibiotic production (bioassay, LC) A B C D E F G H I K L M mg ml-1 Which values will be most similar (small SD, standard deviation)? Which measurements represent independent repeats? The 3 mutant strains should be the same; why could we get very different measurements?

13. All the values in Group 1 are dependent on each other because they all go back to mutant 1 • Only measurements from Group 1 measurements from Group 2 qualify as independent repeats Group 1 Group 2 • The values from Group 1 and Group 2 could be very different if one of them had suffered an additional spontaneousmutation. • Additional spontaneous mutations are common if a gene replacement causes the cells to grow slowly because Compensatory mutations may make the strain fitter.

14. Streptomyces coelicolor Mutants

15. How to prove that the mutation you created causes the observed phenotype? Genome Wild-type gene

16. Introducing a specific mutation into the genome by gene replacement Genome Planned, site-directed mutation The wild-type gene was replaced by a mutant gene or deleted completely by replacement e.g. with a selectable antibiotic resistance gene

17. How to confirm that the observed phenotype was caused the mutation you created, and not by a surprise spontaneous mutation somewhere else in the genome ? Genome I there another spontaneous mutation? Planned, site-directed mutation • What experiment(s) could you do to find out? • explain on paper using a diagram with minimal text.

20. How to confirm that the observed phenotype was caused the mutation you created, and not by a surprise spontaneous mutation somewhere else in the genome ? Complement the mutationin-trans with a copy of the wild-type gene Genome I there another spontaneous mutation? Planned, site-directed mutation Wild-type gene in-trans means ‘somewhere else’ ciscomplementation means repairing the mutated gene itself Plasmid

21. Interpretation of experiments Contamination of cell cultures Independent repeats Spontaneous mutations trans complementation