分子生物学（双语）3 Methods in Molecular Biology and Genetic Engineering.ppt
Chapter 3 Methods in Molecular Biology and Genetic Engineering FIGURE CO: Methods in Molecular Biology and Genetic Engineering © Nicemonkey/Dreamstime.com FIGURE 01: The target of a phosphatase (a) and a nuclease (b). An endonuclease (c) and an exonuclease (d) FIGURE 02: Restriction endonuclease FIGURE 03: A restriction map is a linear sequence of sites separated by defined distances on DNA FIGURE 04: Plasmid transformation FIGURE 05: E. coli colonies on agar plates with ampicillin, IPTG, and the color indicator X-gal FIGURE 06: Several types of cloning vectors are available FIGURE 07: A vector can be used in both yeast and bacteria FIGURE 08: Luciferase graph/Firefly Photo © Cathy Keifer/Dreamstime.com FIGURE 09: A mouse promoter controls tissue-specific expression of lacZ Photo courtesy of Robb Krumlauf, Stowers Institute for Medical Research FIGURE 10A: Fluorescent proteins are powerful research tools Courtesy of Joachim Goedhart, Molecular Cytology, SILS, University of Amsterdam. FIGURE 10B: Fluorescent proteins are powerful research tools Reprinted from Vision Res., vol. 45, T. G. Wensel, et al., Rhodopsin-EGFP knock-ins., pp. 3445-3453. Copyright 2005, with permission from Elsevier http:/www.sciencedirect.com/science/journal/00426989. Photo courtesy of Theodore G. Wensel, Baylor Col FIGURE 11: DNA can be introduced into cells in several ways FIGURE 12: An autoradiogram of a gel prepared from the colonies described in Figure 3.5 FIGURE 13: The fluorescent in situ hybridization (FISH) technique Adapted from an illustration by Darryl Leja, National Human Genome Research Institute (www.genome.gov). FIGURE 14a b: DNA sizes can be determined by gel electrophoresis Adapted from an illustration by Michael Blaber, Florida State University. FIGURE 15: Agarose gel electrophoresis pattern of SV40 DNA Reproduced from W. Keller, Proc. Natl. Acad. Sci. USA 72 (1975): 2550- 2554. Photo courtesy of Walter Keller, University of Basel. FIGURE 16: Gradient centrifugation separates samples based on their density FIGURE 17: DideoxyNTP sequencing using fluorescent tags Inset photo courtesy of Jan Kieleczawa FIGURE 18: Creating a complementary copy of one template strand through denaturation and rapid cooling in the presence of excess primer FIGURE 19: Thermally driven cycles of primer extension lead to exponential production of the short, primer-to-primer defined sequence (the “amplicon”) FIGURE 20: Fluorescence Resonant Energy Transfer (FRET) FIGURE 21: Southern blot FIGURE 22: Poly(A)+ RNA can be separated from other RNAs by fractionation on an oligo(dT) column FIGURE 23: Western blot FIGURE 24: Microarrays show the levels of all the expressed genes in an experimental sample. FIGURE 25: Chromatin immunoprecipitation detects protein-DNA interactions in the native chromatin context in vivo FIGURE 26: Transfected DNA can be incorporated into the mouse genome Photo reproduced from P. Chambon, Sci. Am. 244 (1981): 60-71. Used with permission of Pierre Chambon, Institute of Genetics and Molecular and Cellular Biology, College of France. FIGURE 27: A transgene can cure a disease FIGURE 28: ES cells can be used to generate mice FIGURE 29: Transgenes can be selected FIGURE 30: Cre excises the sequence between lox sites Structure from Protein Data Bank: 1OUQ. E. Ennifar, et al., Nucleic Acids Res. 31 (2003): 5449-5460. FIGURE 31: Cre/lox excises a target only when Cre is activated FIGURE 32: A knock-in replaces an endogenous gene with an alternative sequence
