sign in sign up
<<
Home , Molecular biology

Molecular of Photosynthesis of Photosynthesis

Edited by

GOVINDJEE, Editor-in-Chief University ofIllinois at Urbana, U.sA.

Editorial Board

Hans J. BOHNERT University ofArizona at Tucson, U.S A. w. BOTTOMLEY CSIRO at Canberra City, Australia

Donald A. BRYANT Pennsylvania State University at University Park, U.S A

John E. MULLET Texas A & M University at College Station, U.S A.

W.L. OGREN USDAIARS, University ofIllinois at Urbana, U.S A.

Himadri PAKRASI Washington State University at St. Louis, U.SA.

c.R. SOMERVILLE Michigan State University at East Lansing, U.S.A.

Reprinted from Photosynthesis Research, Vols. 16 - 19, 1988 -1989.

Kluwer Academic Publishers DORDRECHT/BOSTON/LONDON of Congress Cataloging in Publication Data

Molecular biology of photosynthesis / edited by Govindjee. p. cm. Reprinted from Photosynthesis research. Inc 1 udes index. 1. Photosynthesis. 2. molecular . 1. Govindjee, 1933- OK882.M833 1989 581.1·3342--dc19 88-34046 CIP ISBN-13: 978-94-010-7517-6 e-ISBN-13: 978-94-009-2269-3 DOl: 10.1007/978-94-009-2269-3

Published by Kluwer Academic Publishers, P.O. Box 17, 3300 AA Dordrecht, The Netherlands.

Kluwer Academic Publishers incorporates the publishing programmes of D. Reidel, Martinus Nijhoff, Dr W. Junk and MTP Press.

Sold and distributed in the U.S.A. and Canada by Kluwer Academic Publishers, 101 Philip Drive, Norwell, MA 02061, U.S.A.

In all other countries, sold and distributed by Kluwer Academic Publishers Group, P.O. Box 322, 3300 AH Dordrecht, The Netherlands.

All Rights Reserved © 1988 by Kluwer Academic Publishers Softcover reprint of the hardcover I st edition 1988 No part of the material protected by this copyright notice may be reproduced or utilized in any form or by any means, electronic or mechanical including photocopying, recording or by any information storage and retrieval system, without written permission from the copyright owner. Preface

Molecular biology, particularly , is among the newest and most powerful approach in modern photosynthesis research. Development of molecular biology techniques has provided new methods to solve old problems in many biological disciplines. Molecular biology has its greatest potential for contribution when applied in combination with other disciplines, to focus not just on and , but on the complex between them and the biochemical pathways in the whole . Photosynthesis is surely the best studied research area in plant biology, making this field the foremost candidate for successfully employing molecular genetic techniques. Already, the success of molecular biology in photosynthesis has been nothing short of spectacular. Work performed over the last few years, much of which is sum• marized in this volume, stands in evidence. Techniques such as site-specific mutagenesis have helped us in examining the roles of individual domains in the of multiunit complexes such as the ribulose-l ,5-bisphos• phate carboxylase/oxygenase (RUBISCO) and the oxygen evolving photo• system (the photosystem II). The techniques of molecular biology have been very important in advancing the state of knowledge of the reaction center from the photosynthetic whose has been elegantly deduced by H. Michel and 1. Deisenhofer from the X-ray studies of its crystals. Molecular biology can significantly aid in the design of new experiments in photosynthesis research, experiments that can lead to the verification of mech• anisms, to the design of new hypotheses which subsequently may be tested by genetic/protein engineering and studies of engineered . Considering the wealth of our knowledge on how chemical energy is generated from light energy (see books edited by 1. Amesz, 1. Barber and Govindjee during the last several years), it seems that molecular biology has come of age at the most appropriate time for its full exploitation in understanding the molecular mech• anisms of photosynthesis. Problems that have plagued our thoughts for decades are beginning to yield to these powerful techniques. One specific example of the successful application of molecular genetics to photosynthesis has been the recent chemical identifica• tion of an intermediate labeled "D" (or "F"), that acts as a slow electron donor to the reaction center chlorophyll a P680, as a tyrosine on the D2 protein rather than the previously presumed plastoquinol (Debus et al.; Vermaas et al.). This has allowed the suggestion that the normal electron donor "Z" may also be a tyrosine . Furthermore, the amino acid similarity between the "L" and "M" subunits of the bacterial reaction centers and the D J and D2 subunits of plant photosystem II has led to suggestions as to the binding sites of the various chromophores and of the herbicides in the plant photosystem II. In contrast, differences have led to suggestions as to the binding sites of the components of VI

the oxygen evolving complex on the lumenal side of 0, and O 2 since only , not photosynthetic bacteria, evolve oxygen. The of genes and examin• ation of control of expression has led to information on the translational control of the synthesis of chlorophyll-binding when chlorophyll is not synthesized. Additionally, use of gene-fusion techniques has enabled us to raise specific against individual protein components and to use them to advance functional studies. We suspect that each of you will have a personal list of important problems to which molecular genetics has contributed long• awaited insights. There is a much wider and continuing technological revolution in the molecu• lar biology of photosynthesis. The entire nucleotide sequence of the chloroplast of two plants (tobacco and the liverwort [Marchantia polymorpha]) is now available. Several other complete sequences will inevitably be generated. The publication of this book, a compilation of four special issues of the journal Photosynthesis Research, "Molecular Biology of Photosynthesis", marks the achievements of this field. The book is unique in capturing this current excite• ment in its entirety. Twenty-eight authoritative mini reviews and ten original regular papers on this and related topics are included in this book. Topics range from the and the composition of the genes, molecular genetic methods, use of molecular biology in understanding the antenna system (the light harvest• ing system), the reaction centers, herbicide resistance, photosystem I, photo• system II, bacterial system, electron transport complexes and components, A TP synthase, carbon fixation, RUBISCO, Crassulacean Acid (CAM), , and transport of proteins. We hope that a collection of these "burning" topics in photosynthesis research, as assembled in this volume, will be of great use to students of agriculture, , , , plant biology, plant physi• ology, and molecular biology. Several articles contain discussions of newer techniques. Technical terms, which are frequently used in this field, as well as a subject index, have been included to increase the usefulness of this book. We feel that a rapid transfer of technology from the area of basic molecular genetics to the functional studies on photosynthesis is currently taking place. We are fortunate to be part of this exciting revolution in the field of photosynthesis. This is, however, only a beginning and much more remains to be done. We are convinced that for years to come, the continued development of molecular biology as a research tool will be an important contributor in our efforts to thoroughly understand how plants convert the solar energy into chemical energy. The editors of this book are grateful to the chief editors Drs. R. Marcelle and R. Blankenship and all the associate editors of Photosynthesis Research, Mr. Ir. A.C.Plaizier, Ms. Josje Dominicus and Ms. Maria Vermeulen-James of Kluwer Publishers for their cooperation and help during the publication of this book. We also thank Dr. John C. Cushman (University of Arizona) for the glossary of terms and Dr. Rajni Govindjee (University of Illinois) for the subject index.

EDITORS Contents

Preface v Contributors Xl Glossary of terms John C. Cushman XVll

I. Genes and genetics 1. Nicotiana chloroplast genes for components of the photosyn- thetic apparatus. by Kazuo Shinozaki, Nobuaki Hayashida and Masahiro Sug- iura 2. Gene organization and newly identified groups of genes of the chloroplast genome from a liverwort Marchantia polymorpha. by Kanji Ohyama, Takayuki Kohchi, Hideya Fukuzawa, Tohru Sano, Kazuhiko Umesono and Haruo Ozeki 27 3. in chloroplast protein-coding genes of land plants. by Aine L. Plant and John C. Gray 43 4. Pea chloroplast tRNA Lys (UUU) gene: and analysis of an -containing gene. by Scott K. Boyer and John E. Mullet 61 5. Transposon mutagenesis of nuclear photosynthetic genes in Zea mays. by William B. Cook and Donald Miles 77 6. Using bacteria to analyze sequences involved in chloroplast . by Anthony A. Gatenby, Steven J. Rothstein and Douglas Bradley 105 7. Binding, uptake and expression of foreign DNA by cyanobac- teria and isolated etioplasts. by Bruce A. McFadden and Henry Daniell 121

II. Light harvesting systems 8. The puf regoin of Rhodobacter sphaeroides. by Timothy J. Donohue, Patricia J. Kiley and Samuel Kaplan 137 viii

9. Characterization of phycobiliprotein and linker polypeptide genes in Fremyella diplosiphon and their regulated expression during complementary chromatic . by Arthur R. Grossman, Peggy G. Lemaux, Pamela B. Conley, Brigitte U. Bruns and Lamont K. Anderson 161 10. Photoregulation of gene expression in the filamentous cyano- bacterium Calothrix sp. PCC 7601: light-harvesting complexes and cell differentiation. by Nicole Tandeau De Marsac, Didier Mazel, Thierry Damerval, Gerard Guglielmi, Veronique Capuano and Jean Houmard 195 II. Regulation of cyanobacterial -protein composition and organization by environmental factors. by Harold Riethman, George Bullerjahn, K.J. Reddy and Louis A. Sherman 229 12. The major light-harvesting complex of Photosystem II: aspects of its molecular and . by Parag R. Chitnis and J. Philip Thornber 259 13. Regulation and expression of the multigene family coding light- harvesting chlorophyll alb binding proteins of Photosystem II. by Dennis E. Buetow, Houqi Chen, Geza Erdos and Lee S.H. Yi 283

1lI. Reaction centers; herbicide resistance 14. Reaction centers from three herbicide-resistant mutants of Rhodobacter sphaeroides 2.4.1: and preliminary characterization. by Mark L. Paddock, Scott H. Rongey, Edward C. Abresch, George Feher and Melvin Y. Okamura 321 15. Molecular genetics of herbicide resistance in cyanobacteria. by Judy Brusslan and Robert Haselkorn 343 16. of two new resulting in herbicide re- sistance in the cyanobacterium Synechococcus sp. PCC 7002. by Jeffrey C. Gingrich, Jeffrey S. Buzby, Veronica L. Stirewalt and Donald A. Bryant 353 17. Nucleotide sequence of the genes encoding cytochrome b-559 from the cyanelle genome of Cyanophora paradoxa. by Amanda Cantrell and Donald A. Bryant 371 18. Protein composition of the Photosystem II core complex in genetically engineered mutants of the cyanobacterium Syne- chocystis sp. PCC 6803. by W.F.J. Vermaas, M. Ikeuchi and Y. Inoue 389 IX

19. The QB site modulates the conformation of the Photosystem II reaction center polypeptides. by Achim Trebst, Brigitte Depka, Bernd Kraft and Udo Johanningmeier 407 20. Photoregulation of psbA transcript levels in mustard cotyledons. by J.E. Hughes and G. Link 423 21. The molecular mechanism of the Bicarbonate effect at the Plastoquinone reductase site of Photosynthesis. by Danny J. Blubaugh and Govindjee 441 22. Photosystem I complex. by Patricia Reilly and Nathan Nelson 485

IV. Electron transport components; ATP synthase 23. Synthesis and assembly of the cytochrome b-f complex in higher plants. by David L. Willey and John C. Gray 497 24. Genes encoding ferredoxins from Anabaena sp. PCC 7937 and Synechococcus sp. PCC 7942: structure and regulation. by Jan Van Der Plas, Rolf De Groot, Martin Woortman, Fons Cremers, Mies Borrias, Gerard Van Arkel and Peter Weisbeek 517 25. Structure, organization and expression of cyanobacterial A TP synthase genes. by Stephanie E. Curtis 543 26. The chloroplast genes encoding subunits of the H+ -ATP syn- thase. by Graham S. Hudson and John G. Mason 565

V. Carbon; Rubisco 27. Structural gene regions of Rhodobacter sphaeroides involved in

CO2 fixation. by Paul L. Hallenbeck and Samuel Kaplan 583 28. Uptake and utilization of inorganic carbon by cyanobacteria. by John Pierce and Tatsuo Ornata 593 29. Synthesis and assembly of bacterial and higher plant Rubisco subunits in Escherichia coli. by Anthony A. Gatenby 607 30. Organization and expression of the genes encoding ribulose-l ,5- bisphosphate carboxylase in higher plants. by Thianda Manzara and Wilhelm Gruissem 621 31. Cloning, expression and directed mutagenesis of the genes for ribulose bisphosphate carboxylase/oxygenase. by Bruce A. McFadden and Christopher L. Small 645 x

32. The Rubisco subunit binding protein. by R. John Ellis and Saskia M. Van Der Vies 661 33. The value of mutants unable to carry out photorespiration. by Ray D. Blackwell, Alan J.S. Murray, Peter J. Lea, Alan C. Kendall, Nigel P. Hall, Janice C. Turner and Roger C. Wallsgrove 677 34. Gene expression during CAM induction under salt stress in Mesembryanthemum: cDNA library and increased levels of mRNA for phosphoenolpyruvate carboxylase and pyruvate orthophosphate dikinase. by Jurgen M. Schmitt, Christine Michalowski and Hans J. Bohnert 699

VI. Protein transport; synthesis

35. Transport of proteins into chloroplasts. by Thomas H. Lubben, Steven M. Theg and Kenneth Keegstra 713 36. Protein transport towards the thylakoid lumen: post-- al translocation in tandem. by Sjef Smeekens and Peter Weisbeek 735 37. Recent developments in chloroplast transport. by Michael L. Mishkind and Scott E. Scioli 745 38. Protein synthesis by isolated chloroplasts. by A. Gnanam, C.C. Subbaiah and R. Mannar Mannan 777

Subject index 801 Contributors

Abresch, Edward c., Department of Physics B-019, University of California at San Diego, LaJolla CA 92093, USA (pp 321-342) Anderson, Lamont K., Carnegie Institution of Washington, 290 Panama Street, Stanford CA 94305, USA (pp 161-194) Blackwell, Ray D., Department of Biological Sciences, University of Lancaster, Lancaster LA1 4YQ, UK (pp 677-698) Blubaugh, Danny J., Department of Agronomy, N-212 Ag. Sci. N., University of Kentucky, Lexington KY 40546, USA (pp 441-484) Bohnert, Hans J., Biochemistry Department, University of Arizona, Tucson AZ 85721, USA (pp 699-711) Borrias, Mies, Department of Molecular Cell Biology and Institute of Molecular Biology, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Nether• lands (pp 517-542) Boyer, Scott K., Biological Sciences Department, Purdue University, Lilly Hall of Sciences, West Lafayette IN 47907, USA (pp 43-59) Bradley, Douglas, Department of Molecular Genetics, Institute, Trumpington, Cambridge CB22LQ, UK (pp 105-120) Bruns, Brigitte D., Carnegie Institution of Washington, 290 Panama Street, Stanford CA 94305, USA (pp 161-194) Brusslan, Judy, Department of Molecular Genetics and Cell Biology, University of Chicago, Chicago IL 60637, USA (pp 343-352) Bryant, Donald A., Department of Molecular and Cell Biology, S-101 Frear Building, Penn State University, University Park PA 16802, USA (pp 353-369 & 371-387) Buetow, Dennis E., Department of and Biophysics, University of Illinois at Urbana-Champaign, 407 South Goodwin Avenue - 524 Burrill Hall, Urbana IL 61801, USA (pp 283-319) Bullerjahn George, Division of Biological Sciences, University of Missouri, Tucker Hall, Columbia MO 65211, USA (pp 229-257) Buzby, Jeffrey S., Department of Biology, University of California, Los Angeles CA 90024, USA (pp 353-369) Cantrell, Amanda, Department of Molecular and Cell Biology, S-101 Frear Building, Penn State University, University Park PA 16802, USA (pp 371- 387) Capuano, Veronique, Department of Molecular and Cellular Biology, Penn State Unitersity, University Park PA 16802, USA (pp 195-228) Chen, Houqi, Department of Physiology and Biophysics, University of Illinois at Urbana-Champaign, 407 South Goodwin Avenue-524 Burrill Hall, Urbana IL 61801, USA (pp 283-319) xii

Chitnis, Parag R., Biology Department and Molecular Biology Institute, University of California, Los Angeles CA 90024, USA (pp 259-281) Conley, Pamela B., Carnegie Institution of Washington, 290 Panama Street, Stanford CA 94305, USA (pp 161-194) Cook, William B., Department of Biological Sciences, 108 Tucker Hall, Univer• sity of Missouri, Columbia MO 652Jl, USA (pp 77-103) Cremers, Fons, Department of Molecular Cell Biology and Institute of Molecular Biology, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Nether• lands (pp 517-542) Curtis, Stephanie E., Department of Genetics, Box 7614, North Carolina State University, Raleigh NC 27695-7614, USA (pp 543-564) Cushman John c., Biochemistry Department, University of Arizona, Tucson AZ 85721, USA Damerval, Thierry, Departement de Biochimie et Genetique Mo!eculaire, Unite de Physiologie Microbienne (CNRS UA 1129), Institut Pasteur, 28, rue du Docteur Roux, 75724 Paris Cedex 15, France (pp 195-228) Daniell, Henry, Biochemistry/Biophysics Program, Washington State University, Pullman WA 99164-4660, USA (pp 121-135) De Groot, Rolf, Department of Molecular Cell Biology and Insitute of Molecular Biology, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands (pp 517-542) Depka, Brigitte, Department of Biology, Ruhr-University of Bochum, P.o. Box 102148, D-4630 Bochum 1, FRG (pp 407-421) Donohue, Timothy J., Bacteriology Department, University of Wisconsin, 1550 Linden Drive, Madison WA 53706, USA (pp 137-159) Ellis, R. John, Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK(pp 661-675) Erdos, Geza, Department of Physiology and Biophysics, University of Illinois at Urbana-Champaign, 407 South Goodwin Avenue - 524 Burrill Hall, Urbana IL 61801, USA (pp 283-319) Feher, George, Department of Physics B-019, University of California at San Diego, LaJolla CA 92093, USA (pp 321-342) Fukuzawa, Hideya, Institute of Applied , University of Tokyo, Tokyo 113, Japan (pp 27-42) Gatenby, Anthony A., Central Research and Development Department, Experimental Station, E.l. DuPont de Nemours & Co., Wilmington DE 19898, USA (pp 105-120 & 607-619) Gingrich, Jeffrey c., Chemical Biodynamics Division, Lawrence Berkely Laboratory, University of California, Berkeley CA 94720, USA (pp 353- 369) Gnanam, A., Department of Plant Sciences, School of Biological Sciences, Madurai Kamara} University, Madurai 625021, India (pp 777-800) Govindjee, Departments of Plant Biology, Physiology and Biophysics, Xlll

University of Illinois at Urbana-Champaign, 505 South Goodwin Avenue- 289 Morrill Hall, Urbana IL 61801, USA (pp 441-484) Gray, John c., School, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK (pp 43-59 & 497-516) Grossman, Arthur G., Carnegie Institution of Washington, 290 Panama Street, Stanford CA 94305, USA (pp 161-194) Gruissem, Wilhelm, Department of Botany, University of California, Berkeley CA 94720, USA (pp 621-644) Guglielmi, Gerard, Departement de Biochimie et Genhique Moleculaire, Unite de Physiologie Microbienne (CNRS U A 1129), Institut Pasteur, 28, rue du Docteur Roux, 75724 Paris Cedex 15, France (pp 195-228) Hall, Nigel P., Biochemistry Department, Rothamsted Experimental Station. Harpenden Herts AL5 2JQ, UK (pp 677-698) Hallenbeck, Paul L., Department of Microbiology, University of Illinois, 407 South Goodwin Avenue-136 Burrill Hall, Urbana IL 61801, USA (pp 583-591) Haselkorn, Robert, Department of Molecular Genetics and Cell Biology, University of Chicago, Chicago IL 60637, USA (pp 343-352) Hayashida, Nobuaki, Center for Gene Research, Nagoya University, Chikusa, Nagoya 464, Japan (pp 1-25) Houmard, Jean, Departement de Biochimie et Genhique Moleculaire, Unite de Physiologie Microbienne (CNRS UA 1129), Instilut Pasteur, 28, rue du Docteur Roux, 75724 Paris Cedex 15, France (pp 195-228) Hudson Graham S., Division of Plant Industry, CSIRO, GPO Box 1600, Canberra, A.C.T. 2601, Australia (pp 565-582) Hughes, J.E., Department of Botany, University of Georgia, Athens GA 30602, USA (pp 423-439) Ikeuchi, M., Solar Energy Research Group, RIKEN, Hirosawa 2-1, Wako• shi, SaUama 351-01, Japan (pp 389-405) Inoue, Y., Solar Energy Research Group, RIKEN, Hirosawa 2-1, Wako-shi, Saitama 351-01, Japan (pp 389-405) Johanningmeier, Udo, Department of Biology, Ruhr-University of Bochum, P.O. Box 102148, D-4630 Bochum 1, FRG (pp 407-421) Kaplan, Samuel, Department of Microbiology, University of Illinois at Ur• bana-Champaign, 407 South Goodwin Avenue-136 Burrill Hall, Urbana IL 61801, USA (pp 137-159 & 583-591) Keegstra, Kenneth, Botany Department, University of Wisconsin, Madison WI 53706, USA (pp 713-734) Kendall, Alan c., Biochemistry Department, Rothamsted Experimental Station, Harpenden, Herts AL5 2JQ, UK (pp 677-698) xiv

Kiley, Patricia J., Microbiology Department, University of Illinois, 407 South Goodwin Avenue-136 Burrill Hall, Urbana IL 61801, USA (pp 137-159) Kohchi, Takayuki, Research Center for Cell and Culture, Faculty of Agriculture, Kyoto University, Kyoto 606, Japan (pp 27-42) Kraft, Bernd, Department of Biology, Ruhr-University of Bochum, P.O. Box 102148, D-4630 Bochum 1, FRG (pp 407-421) Lea, Peter J., Department of Biological Sciences, University of Lancaster, Lancaster LA1 4YQ, UK (pp 677-698) Lemaux, Peggy G., Carnegie Institution of Washington, 290 Panama Street, Stanford CA 94305, USA (pp 161-194) Link, G., Pjlanzliche Zellphysiologie, Ruhr-University of Bochum, P.o. Box 10 21 48, D-4630 Bochum 1, FRG (pp 423-439) Lubben, Thomas H., Botany Department, University of Wisconsin, Madison WI 53706, USA (pp 713-734) Mannan, R. Mannar, Department of Plant Sciences, School of Biological Sciences, Madurai Kamara} University, Madurai 625021, India (pp 777- 800) Manzara, Thianda, Department of Botany, University of California, Berkeley CA 94720, USA (pp 621-644) Mason, John G., Division of Plant Industry, CSIRO, GPO Box 1600, Canberra, A.C.T. 2601, Australia (pp 565-582) Mazel, Didier Departement de Biochimie et Genetique Moleculaire, Unite de Physiologie Microbiimne (CNRS UA 1129), Institut Pasteur, 28, rue du Docteur Roux, 75724 Paris CMex 15, France (pp 195-228) McFadden, Bruce A., Biochemistry/Biophysics Program, Washington State University, Pullman WA 99164-4660, USA (pp 121-135 & 645-660) Michalowski, Christine, Biochemistry Department, University of Arizona, Tucson AZ 85721, USA (pp 699-711) Miles; Donald, Department of Biological Sciences, 108 Tucker Hall, Univer• sity of Missouri, Columbia MO 65211, USA (pp 77-103) Mishkind, Michel L., Department of Biochemistry and Microbiology, Lipman Hall, Cook College, Rutgers University, New Brunswick NJ 08903, USA (pp 745-776) Mullet, John E., Department of Biochemistry & Biophysics, Texas A & M University, College Station TX 77843-2128, USA (pp 43-59) Murray, Alan J.S., Department of Biological Sciences, University of Lan• caster, Lancaster LA1 4YQ, UK (pp 677-698) Nelson, Nathan, Roche Institute of Molecular Biology, Roche Research Center, Nutley NJ 07110, USA (pp 485-496) Ohyama, Kanji, Research Center for Cell and , Faculty of Agriculture, Kyoto University, Kyoto 606, Japan (pp 27-42) xv

Okamura, Melvin Y., Department of Physics B-019, University of California at San Diego, LaJolla CA 92093, USA (pp 321-342) Omata, Tatsuo, Solar Energy Research Group, The Institute of Physical and Chemical Research (RIKEN) , Wako-shi, Saitama, 35/-01, Japan (pp 593-606) Ozeki, Hamo, Department of Biophysics, Faculty of Science, Kyoto Univer• sity, Kyoto 606, Japan (pp 27-42) Paddock, Mark L., Department of Physics B-OJ9, University of California at San Diego, LaJolla CA 92093, USA (pp 321-342) Pierce, John, Central Research and Development Department, E.I. DuPont de Nemours and Co., Experimental Station, Building 402, Room 2230, Wil• mington DE 19898, USA (pp 593-606) Plant, Aine L., Botany School, University of Cambridge, Downing Street, Cambridge CB23EA, UK (pp 43-59) Reddy, K.J., Division of Biological Sciences, University of Missouri, Tucker Hall, Columbia MO 65211, USA (pp 229-257) Reilly, Patricia, Roche Institute of Molecular Biology, Roche Research Center, Nutley NJ 07110, USA (pp 485-496) Riethman, Harold, Division of Biological Sciences, University of Missouri, Tucker Hall, Columbia MO 65211, USA (pp 229-257) Rongey, Scott H., Department of Physics B-OJ9, University of California at San Diego, LaJolla CA 92093, USA (pp 321-342) Rothstein, Steven J., Department of Molecular Biology & Genetics, Univer• sity of Guelph, Ontario NIG 2Wl, Canada (pp 105-120) Sano, Tohm, Research Center for Cell and Tissue Culture, Faculty of Agri• culture, Kyoto University, Kyoto 606, Japan (pp 27-42) Schmitt, Jurgen M., Botanisches Institut der Universitat Wurzburg, 8700 Wiirzburg, FRG (pp 699-711) Scioli, Scott E., Department of Biochemistry and Microbiology, Lipman Hall, Cook College, Rutgers University, New Brunswick NJ 08903, USA (pp 745-776) Sherman, Louis A., Division of Biological Sciences, University of Missouri, Tucker Hall, Columbia MO 65211, USA (pp 229-257) Shinozaki, Kazuo, Center for Gene Research, Nagoya University, Chikusa, Nagoya 464, Japan (pp 1-25) Small, Christopher L., Biochemistry/Biophysics Program, Washington State University, Pullman WA 99164-4660, USA (pp 645-660) Smeekens, Sjef, Department of Molecular Cell Biology and Institute of Mole• cular Biology, University of Utrecht, Padualaan 8, 3584 CH Utrecht, The Netherlands (pp 735-744) Stirewalt, Veronica L., Department of Molecular and Cell Biology, S-lOJ XVI

Frear Building, Penn State University, University Park PA 16802, USA (pp 353-369) Subbaiah, e.e., Department of Plant Sciences, School of Biological Sciences, Madurai Kamaraj University, Madurai 625021, India (pp 777-800) Sugiura, Masahiro, Center for Gene Research, Nagoya University, Chikusa, Nagoya 464, Japan (pp 1-25) Tandeau De Marsac, Nicole, Departement de Biochimie et Genetique MoJe• culaire, Unite de Physiologie Microbiimne (CNRS UA 1129), Institut Pasteur, 28, rue du Docteur Roux, 75724 Paris CMex Roux, France (pp 195-228) Theg, Steven M., Department of Botany, University of California at Davis, Davis CA 95616, USA (pp 713-734) Thornber, J. Philip, Biology Department and Molecular Biology Institute, University of Calfornia, Los Angeles CA 90024, USA (pp 259-281) Trebst, Achim, Department of Biology, Ruhr-University of Bochum, P.o. Box 102148, D-4630 Bochum 1, FRG (pp 407-421) Turner, Janice e., Biochemistry Department, Rothamsted Experimental Station, Harpenden, Herts AL5 2JQ, UK (pp 677-698) Umesono, Kazuhiko, Department of Biophysics, Faculty of Science, Kyoto University, Kyoto 606, Japan (pp 27-42) Van Arkel, Gerard, Department of Molecular Cell Biology and Institute of Molecular Biology, University of Utrecht, Padualaan 8,3584 CH Utrecht, The Netherlands (pp 517-542) Van Der Plas, Jan, Department of Molecular Cell Biology and Institute of Molecular Biology, University of Utrecht, Padualaan 8,3584 CH Utrecht, The Netherland!} (pp 517-542) Van der Vies, Saskia M., Department of Biological Sciences, University of Warwick, Coventry CV4 7AL, UK (pp 661-675) Vermaas, W.F.J., Department of Botany, Arizona State University, Tempe AZ 85787-1601, USA (pp 389-405) Walls grove, Roger M., Biochemistry Department, Rothamsted Experimental Station, Harpenden, Herts AL5 2JQ, UK (pp 677-698) Weisbeek, Peter, Department of Molecular Cell Biology and Institute of Molecular Biology, University of Utrecht, Padualaan 8,3584 CH Utrecht, The Netherlands (pp 517-542 & 735-744) Willey, David L., Botany School, University of Cambridge, Downing Street, Cambridge CB2 3EA, UK (pp 497-516) Woortman, Martin, Department of Molecular Cell Biology and Institute of Molecular Biology, University of Utrecht, Padualaan 8,3584 CH Utrecht, The Netherlands (pp 517-542) Yi, Lee S.H., Department of Physiology and Biophysics, University of Illinois at Urbana-Champaign, 407 South Goodwin Avenue-524 Burrill Hall, Urbana IL 61801, USA (pp 283-319) Govindjee et aI. (eds), Molecular Biology of Photosynthesis: xvii © 1988 Kluwer Academic Publishers

Glossary of terms

Compiled by: John C. Cushman Department of Biochemistry, University of Arizona, Tucson, Arizona 85721, USA

Adapter Small synthetic which when annealed and ligated to blunt ended DNA molecules from "Preformed" protruding termini which do not require cleavage with a to create a cohesive end (see linker). gel A technique used to separate according to size, to identify and purify DNA fragments. Used in conjunction with Southern blotting and Northern blotting (for RNA). (bacterial or calf intestina!) An enzyme that catalyses the removal of 5' phosphate residues from DNA or RNA. Used in constructing recombinant molecules to prevent circularization of vectors without inserts. Amplification The process of propagating a primary or secondary library created in or bacteriophage (lambda) vectors which greatly increases the number of clones. Amplification may also refer to the production of additional copies of a chromosomal sequence. Anticodon Trinucleotide sequence present on tRNA molecules that is complemen• tary to the codon representing a specific amino acid. ATA Aurintricarboxylic Acid. Used as a potent inhibitor of RNAses during preparation of RNA. Bacteriophage (phage) A that infects and may kill bacterial cells. See lambda phage and M13. Bal31 nuclease An enzyme that catalyses the removal of nucleotides from both 5'- and 3'-termini of double-stranded and single-stranded DNA creating shorter molecules. Used in restriction mapping and in vitro mutagenesis. xviii Blunt end ligation See ligation. Blunt (flush) ends Completely base paired termini of double-stranded DNA molecules. CAAT Box A conserved sequence located about 75 bp upstream from the start point of transcription in . cDNA (complementary DNA) A DNA molecule that is complementary to an RNA molecule. Usually synthesized by the enzyme . cDNA cloning A technique which copies mRNA into DNA using reverse transcriptase followed by the synthesis of a second strand of DNA and insertion of double-strand DNA into a . Chloroplast gene nomenclature Chloroplast genes are named according to the recommendations set forth by: Hallick R.B. and Bottomley, W. 1983. Proposals for the Naming of Chloroplast Genes. Plant Molecular Reporter 1: 38-43. The complex of DNA and protein in the nucleus of a cell or nucleoid of a bacterial cell. Chromosome A discrete unit of the genome consisting of proteins and a very long DNA molecule carrying many genes. A member of a of identical DNA molecules or of genetically identical cells or organisms derived from a single individual by asexual processes. Cloning is the term used to designate the generation and propagation of discrete DNA molecules. Codon A triplet ofnucleotides that specify a particular amino acid or a termina• tion signal (). As initiation codon, usually ATG (AUG in RNA) is used. Termination codons are T AA, TAG and TGA. While codon assignment is, in principle, universal, some mitochondria have specific different codon assignments. Cohesive (sticky) ends Single-stranded termini of double stranded DNA molecules that are complementary to one another. Some restriction endonucleases cut DNA so that cohesive ends are formed, others result in blunt ends. Colony hybridization A technique used to detect the presence of a cloned DNA segment in a bacterial colony (see screening). xix Concatemer Two or more identical sequences joined tandemly head to tail. Cosmids vectors which contain phage lambda DNA sequences (COS sites) allowing the DNA to be packaged in vitro into lambda phage particles. The size of DNA inserts that can be cloned is greater than that of lambda vectors. DNA Ligase (T4) An enzyme that catalyses the fomation of phospho diester bonds between adjacent 3'-OH and 5' phosphate termini in double-stranded DNA. Used to join double-stranded DNA molecules with compatible cohesive termini (sticky ends) or to join blunt-ended double-stranded DNA mole• cules or linkers. (See RNA Ligase (T4).) DNA I (E. coli) Polymerizes DNA (needs primer, template and nucleotides) in a 5'- to 3' -direction. Also contains a 5'- to 3' -exonuclease, and a 3'- to 5' -exonu• clease for proofreading. Used for labeling via nick-translation. DNA polymerase (T4) See T4 DNA polymerase. DNase I An endonuclease isolated from bovine pancreas that breaks double or single stranded DNA. In the presence of Mg+ +, DNase I makes single strand nicks in double-stranded DNA; in the presence of Mn + + DNAse cuts double-strand DNA resulting in fragments of approximately the same size. Used in characterizing nucleosomes, and for making single• strand nicks in DNA prior to nick-translation. End Labelling A technique that introduces radioactively labeled nucleotides (dNTPs) which are usually radioactively labeled using Klenow fragment or T4 DNA polymerase or T4 polynucleotide kinase to transfer phosphate groups to the termini of DNA molecules. A specific DNA sequence element that increases the utilization of a in cis. Enhancers can function independently of location and orientation relative to the promoter. Any segment of an interrupted gene represented in fully processed, mature mRNA (see intron) that is translated into amino acids. Exonuclease III An enzyme that catalyses the stepwise removal of 5' mononucleotides in a 3' to 5' direction from 3' -OH ends of double-stranded DNA. Used for preparing single-stranded DNA probes, and for deleting sequences from xx the ends of DNA fragments in conjunction with a S[ nuclease. Commonly used for the production of overlapping deletion subclones used in sequencing large DNA fragments. Exonuclease VII A processive exonuclease that removes small oligonucleotides from the 3' and 5' ends of single-stranded DNA. A vector that allows the synthesis of RNA or protein from a coding region after engineering to contain promoters or other control elements for efficient synthesis in the particular host organism. Footprinting A technique used to identify regions of DNA bound by specific domains of some proteins by virtue of protection against nuclease attack. Gene A segment of DNA involved in producing a polypeptide chain including a promoter and other regulatory elements. A gene includes regions preceding (5'-leader sequence) and following (3'-trailing sequence) in• dividual coding segments (). Gene Cassette A gene or portions of a gene bordered by suitable restriction sites that can be interchanged to produce a desired vector or gene fusion. Gene Fusion (Chimeric genes) Gene constructs in which different gene cassettes have been combined to produce composite transcriptional units or translational reading frames. Gene nomenclature Bacterial gene nomenclature should follow the conventions set forth by: Demerec, M., E.A. Adelberg, A.l. Clark, and Philip E. Hartman. 1966. A proposal for a Uniform Nomenclature in Bacterial Genetics. Genetics 54: 61-76. Increasingly, this nomenclature is also being used for eukar• yotic genes. Genome The entire complement of chromosomal DNA of a given orgamsm. Genomic clones are derived directly from genomic DNA, not from a cDNA copy. Genomic Southern A technique in which total genomic DNA is digested with a restriction endonuclease, separated by agarose and transferred to filters. See Southern blotting. Guanidinium chloride or isothiocyanate Potent chaotropic agents used to inactivate nucleases and disintegrate cell and nucleoproteins from nucleic acids. Used to isolate intact RNA from tissues that are typically rich in RNase. xxi Hybridization Association by base pairing that occurs between two complementary strands of either DNA and/or RNA. Often used to identify a gene from an organism by using a probe from an identical, analogous gene of another organism. The strength of hybridization depends on nucleotide sequence similarity between the . Hybridoma A cell produced as a result of fusing a myeloma with a lymphocyte which will indefinitely express the immunoglobulins of both parent cells. A that triggers transcription of a gene by binding to a regulator protein. Insert Any piece of DNA that has been introduced into a specific site (restriction site) of a vector. Introns (intervening sequences) Segments of DNA that are transcribed, but that are removed from the mature mRNA. The remaining pieces of RNA (exons) are joined together by splicing. In vitro mutagenesis Modifications made to isolated and purified genes by chemical or physical methods. Some modifications made include deletions, insertions, inver• sions and transpositions. Isoschizomer Different restriction endonuclease (type II) which recognize(s) and cleave(s) the same target sequences. Kb (kilobase) An abbreviation for 1000 base pairs of DNA or 1000 bases of RNA. Klenow fragment Derived from DNA polymerase I of E. coli by treatment with subtilisin (a proteolytic enzyme). Some firms now offer a product that is made by recombinant DNA techniques. Lacks 5' to 3' exonuclease activity of the holoenzyme. Used for Sanger DNA sequencing (see sequencing), filling in the 3' recessed termini (5' overhangs) created by some restriction , and for second-strand cDNA synthesis. Lambda exonuclease Digests DNA in a 5' to 3' direction. Lambda phage A tempelate phage containing about 50 kb of DNA. Derivatives of this phage provide a useful vector for cloning large segments of DNA (up to about 20 kb). Genes, or parts of genes, are used in plasmids (e.g.) for promoter constructions or as and terminators of transcription. xxii

Leader The nontranslated sequence at the 5' end of a transcribed mRNA that precedes the initiator codon. Library A collection of cloned random fragments of DNA representing the entire genome. Sometimes called a gene bank. Both cDNA libraries (a collection of the DNA versions of a group of ) and genomic libraries are commonly encountered. Ligation The formation of a phosphodiester bond covalently linking two adjacent bases separated by a nick in one strand of a DNA duplex. The term blunt-end ligation refers to the reaction wherein two blunt-ended DNA duplex molecules are joined covalently at their ends. Linker Self complementary synthetic oligonucleotides of defined sequence con• taining the cleavage site of one or more restriction endonucleases. After these fragments are blunt-end ligated to DNA molecules, the molecule contains the recognition sequence of a restriction endonuclease and can be subsequently cleaved. Lysogen A bacterium that possesses a repressed prophage integrated into its genome. M13 A filamentous bacteriophage that carries a single strand of DNA whose life cycle includes a double-stranded DNA stage. Used in DNA sequenc• ing and for in vitro mutagenesis. Methylases A number of enzymes that introduce methyl groups into DNA. These enzymes may be used to modify specific sites in DNA and prevent their being cut with certain restriction enzymes. Micrococcal nuclease An endonuclease that cleaves DNA and RNA. In chromatin, micrococcal nuclease preferentially cleaves between nucleosomes. mRNA RNA that serves as a template for protein synthesis. Called 'messenger' RNA. Its sequence may differ from that of the gene from which it was transcribed because of the removal of introns and other processing events. Mung-bean nuclease Enzyme with properties similar to S 1 nuclease, but gentler in its action. Used for converting protruding termini to blunt ends. xxiii

Nick The absence of a phosphodiester bond between two adjacent nucleotides on one strand of duplex DNA. Nick translation Nick translation makes use of the 5' and 3' exonuclease activity of DNA polymerase I to move (,translate') a nick in DNA from one position to another. The polymerase starts synthesis of new strand at the nick, degrading the DNA ahead of it. Nick translation is used to label DNA so that it can be used as a probe. Northern blots A technique for analysis of RNA. The RNA is run out on an agarose or acrylamide gel and then transferred to a suitable filter (nitro• cellulose or nylon membrane). The RNA of interest is then detected by hybridization with an appropriately labeled probe. See "Southern blots". Nucleosome The basic structural subunit of chromatin consisting of about 200 bp of DNA and an octamer of histone proteins. Nucleotide (base) The basic structural unit of nucleic acids. A nucleotide consists of a heterocyclic ring of carbon and nitrogen (nitrogenous base), a five carbon sugar ring (pentose) and a phosphate group. The pentose ring distinguishes DNA and RNA. The pentose ring in DNA is 2-deoxyribose, whereas in RNA it is ribose. The nitrogenous bases are of two types: the pyrimidines (cytosine, uracil and thymine) have six-membraned rings and the purines (adenine and guanine) have fused five and six-membered nngs. DNA or RNA molecules of variable (6 to, usually, less than 100 bases) length that may be synthesized in vitro to serve as linkers, adaptors or probes, or templates for site-directed mutagenesis. Papovaviruses containing small circular . Examples are SV 40 and polyoma. Phagemid A plasmid vector which contains regions of DNA required for the pack• aging of the plasmid as single-stranded DNA by an Ml3 bacteriophage derivative called a 'helper phage'. Plasmid An autonomously replicating, circular, extrachromosomal genetic element. xxiv

Plasmid Nomenclature Cloned DNA fragments contained in a vector can be named in a variety of ways, always preceeded by 'p', denoting either a person (pBR), a laboratory (pEMBL), a function of the cloned DNA (ptac), or combina• tions of those terms usually followed by numbers. gel electrophoresis A simple technique to analyze and purify DNA fragments less than I kb length. Polyacrylamide gels are also used for analyzing proteins and for sequencmg. Addition of poly A to the 3'-end of an RNA molecule by a mechanism that does not involve transcription. Polynucleotide kinase (T4) An enzyme that catalyses transfer of the gamma-phosphate of ATP to the 5' -hydroxyl at 5' -termini of RNA or DNA for sequencing by the Maxam• Gilbert technique. Used for labeling DNA and RNA (with 32p-phosphate labeled rA TP), and for restoring the phosphate removed by alkaline phosphatase. Polysome A string of attached to a single mRNA molecule. Primer In DNA synthesis, a single stranded DNA, often an oligonucleotide (oligomer), that serves as a starting point for polymerization of a second chain. The prlmer base pairs with the template (the other strand) and is extended by a DNA polymerase to form a complementary strand. Probe A gene specific DNA fragment from coding, leader or trailer regions which is usually radioactively labeled and used via hybridization to locate and define another gene or mRNA of unknown character. The term is also used for the specific antibodies used to screen gene libraries (see Screening). Promoter In prokaryotes, a region of DNA that is involved in DNA-dependent RNA polymerase recognition and binding. The promoter regulates the site of initiation of transcription as well as its efficiency. In eukaryotes, promoter sequences do not directly bind RNA polymerase. Instead, they are the site of binding of a complex of proteins that are recognized by the polymerase. Prophage A phage genome covalently integrated as a linear part of the bacterial chromosome. xxv

Pseudogene Inactive, but stable version(s) of a gene derived by from an ancestral active gene. Replisome A multi protein complex containing DNA polymerase and other proteins that assembles at the bacterial replication fork to conduct DNA syn• thesis. Restriction Endonucleases (Type II) Enzymes that recognize a particular sequence (usually 4-8 nucleotides with a twofold axis of symmetry) in double-stranded DNA and cut, in or near that sequence, leaving 5' phosphate and 3' hydroxyl protruding cohesive termini or blunt ends. Used to cut DNA molecules into pieces of defined sizes with specific kinds of ends. Restriction Endonuclease Nomenclature Restriction endonucleases are named according to the proposal by: Smith, o. and Nathans. 1973. A Suggested Nomenclature for Bacterial Host Modification and Restriction Systems and Their Enzymes. J. Mol. Current BioI. 81: 419-423. A listing of all known restriction en• donucleases can be found in: Roberts, R.J. 1988. Restriction Enzymes and Their Isoschizomers. Nucleic Acids Research. Sequences Supple• ments 16: r271-r313. Reverse Transcriptase (RNA-Dependent DNA polymerase) An enzyme that catalyzes the polymerization of DNA 5' to 3' using an oligonucleotide primer or single-stranded RNA as a template (DNA may also be used). Used in the first synthesis of cDNA (first strand). The enzymes also contain a processive 5' and 3' ribonuclease specific for RNA: DNA hybrids. RFLP - Restriction fragment length Term used to describe differences in the DNA restriction pattern that distinguishes several genes in an organism or in individuals of a popula• tion. RFLP is used to monitor the number of genes for one character, the of genes and the linkage between DNA rearrangements and mutant characters. Ribonuclease A This enzyme isolated from bovine pancreas is an endoribonuclease which specifically attacks the 3' -phosphate groups of pyrimidine residues cleaving the 5' -phosphate linkage of adjacent nucleotides. Used to remove RNA from DNA preparations. Ribonuclease Tl This enzyme isolated from Aspergillus oryzae is an endoribonuclease which specifically attacks the 3' -phosphate groups of guanosine residues xxvi

cleaving the 5' -phosphate linkage of adjacent nucleotides. (Used to remove large pieces of RNA from DNA preparations). Ribonucleoprotein particles consisting of two subunits that catalyze the assembly of amino acids into polypeptides using mRNA as a template. RNA ligase (T4) An enzyme that catalyzes the covalent linkage of the 5' -phosphate of one molecule (single-stranded DNA or RNA) with the 3' OH of another. Reported to increase the efficiency of blunt-end ligation of DNA mole• cules using T 4 ligase. RNase A group of enzymes with different specificities for the degradation of RNA molecules. RNasin A commercially available enzyme that is a potent inhibitor of RNase. It can be included in enzymatic reactions and easily extracted with . Sj mapping A technique used to locate the ends of RNA molecules in relation to specific sites (e.g. restriction sites) within the template DNA. Sj nuclease Single-strand specific endonuclease isolated from Aspergillus oryzae. DNA/DNA, DNA/RNA, and RNA/RNA hybrids are resistant to its action at low concentrations of the enzyme. Used in "Sj mapping", RNA protection experiments, and cDNA cloning to remove non-complemen• tary areas of DNA/DNA, DNA/RNA, and RNA/RNA hybrids. Screening The act of searching for and isolating a particular cell under particular conditions confered by a selectable marker or searching for a particular gene using a or probe. Selectable Marker A gene or genes that encodes a protein that confers upon a cell the ability to survive under conditions of selection (e.g. antibiotic resistance genes). Sequenase

A modified form ofT7 DNA polymerase, which possesses greater process• ivity than Klenow fragment, used in Sanger-type sequencing. Sequencing A technique for determining the nucleotide sequence of a specific fragment of DNA or RNA employing Klenow, sequenase, or reverse transcriptase and chain terminating dideoxynucleotides (Sanger method) or chemical modification base specific chemical cleavage reactions (Max• am-Gilbert method). xxvii

Shine-Dalgarno Sequence (Ribosome binding site) A poly purine stretch with the consensus prokaryotic feature AGGAGG located just upstream (usually 7-10 bases) of the AUG initiation codon. This sequence is complementary to the sequence at the 3' -end of 16S rRNA and is involved in the binding of ribosomes to mRNA. Shuttle vector A plasmid that is capable of growing in two or more hosts. Site-directed mutagenesis Construction of altered genes or gene products using methods that allow defined changes at a predetermined position in a gene. Examples are nucleotide exchanges which may lead to amino acid exchanges. SnRNA (small nuclear RNA) Small RNA molecules confined to the nucleus involved in splicing. SnuRPS Small nuclear ribonucleoproteins associated with SnRNA involved in splicing and splicosome formation. Southern blotting A technique originated by E. Southern. DNA is transferred from gels after electrophoresis to filters where specific sequences may be detected using radioactively labeled nucleic acid probes. Splicing The removal of introns and joining of exons in RNA molecules. Splicosome A ribonucleoprotein particle of about 40S consisting of about 40S cons• isting of SnRNAs and 7-10 small basic proteins ("SnuRPS") that cataly• zes the splicing out of introns and splicing together of exon. TATA Box (Hogness box) A conserved A-T rich region located about 25 bp upstream from the transcriptional start site thought to position RNA polymerase II for transcriptional initiation. T4 DNA polymerase An enzyme used to label recessed 3' -termini created by some restriction endonucleases. It possesses a more powerful 3'- to 5' -exonuclease activity than Klenow making. Also possesses the 5'- to 3' -polymerase activity. The natural end of a chromosome. Plasmids containing are ma:intained as one copy per cell. Terminal transferase (Terminal Deoxynucleotidyl Transferase) An enzyme that adds deoxynucleotides to the 3'-OH end of single-stran• ded DNA molecules or double-stranded DNA with protruding 3'-OH termini. Used to add homopolymer tails onto DNA before ligation, and for end-labeling the 3' -ends of DNA fragments. xxviii Trailer The non translated sequence located at the 3'-end of a mRNA following the termination codon. Transacting factor A protein that mediates transcription by binding to promoters, enhancers or other cis-acting regulatory elements. Transcription Synthesis of RNA directed by a DNA template using the enzyme RNA• polymerase. Translation Synthesis of protein by ribosomes directed by mRNA. Transfer RNA (tRNA) A small RNA molecule of75-85 nucleotides that recognizes and becomes covalently attached to only one amino-acid to form an amino acyl-tRNA and contains the anticodon for recognizing the codon present in mRNA that represents the specific amino acid. Uptake of purified viral DNA by (most often) bacterial cells. Transformation Induced uptake of DNA of any kind by cells. Transposase The enzyme activity involved in the insertion of a transposon at a new site. Transposon (Transposable elements) A DNA sequence usually carrying a gene or genes that is able to replicate and insert a copy of itself at a new location in the genome of prokaryotes or eukaryotes. Vector Any plasmid, cosmid or phage into which a foreign piece of DNA (insert) may be ligated to be cloned. Western blotting (Immunoblotting) A technique in which proteins are resolved by polyacrylamide gel electro• phoresis and transferred to a filter and probed with antibodies [identified by binding to a secondary labeled antibody] to identify a specific protein of interest.