Contents
Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
v
Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
xi
PART I IN SITU DETECTION OF RNA MOLECULES USING FISH TECHNIQUES
1 Tagging and Application of RNA Probes for Sequence-Specific
Visualization of RNAs by Fluorescent In Situ Hybridization . . . . . . . . . . . . . . . . .
3
Thomas Dresselhaus and Andrea Bleckmann
2 Quantitative Fluorescence In Situ Hybridization Detection
of Plant mRNAs with Single-Molecule Resolution . . . . . . . . . . . . . . . . . . . . . . . . . . 23
Kun Huang, Mona Batish, Chong Teng, Alex Harkess,
Blake C. Meyers, and Jeffrey L. Caplan
3 Visualization of Endoplasmic Reticulum-Associated mRNA
in Mammalian Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
Jingze J. Wu and Alexander F. Palazzo
4 Simultaneous Detection of mRNA and Protein in S. cerevisiae
by Single-Molecule FISH and Immunofluorescence . . . . . . . . . . . . . . . . . . . . . . . . . 51
Evelina Tutucci and Robert H. Singer
PART II IN VIVO IMAGING OF RNA TRANSPORT AND LOCALIZATION
5 Development and Applications of Fluorogen/Light-Up RNA Aptamer
Pairs for RNA Detection and More . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
Michael Ryckelynck
6 Visualization of Transiently Expressed mRNA in Plants Using MS2 . . . . . . . . . . . 103
Eduardo Jose ´ Pen ˜a and Manfred Heinlein
7 New Generations of MS2 Variants and MCP Fusions to Detect
Single mRNAs in Living Eukaryotic Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
Xavier Pichon, Marie-Ce ´cile Robert, Edouard Bertrand,
Robert H. Singer, and Evelina Tutucci
8 In Vivo Imaging of Mobile mRNAs in Plants Using MS2 . . . . . . . . . . . . . . . . . . . . 145
Kai-Ren Luo, Nien-Chen Huang, and Tien-Shin Yu
9 RNA Imaging with RNase-Inactivated Csy4 in Plants and Filamentous
Fungi. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157
David Burnett, Alexander Lichius, and Jens Tilsner
PART III IMAGING AND ANALYSIS OF RNA UPTAKE AND TRANSPORT
BETWEEN CELLS
10 Utilizing Potato Virus X to Monitor RNA Movement. . . . . . . . . . . . . . . . . . . . . . . 181
Zhiming Yu, Sung Ki Cho, Pengcheng Zhang,
Yiguo Hong, and David J. Hannapel
vii
Preface . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
v
Contributors. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
xi
PART I IN SITU DETECTION OF RNA MOLECULES USING FISH TECHNIQUES
1 Tagging and Application of RNA Probes for Sequence-Specific
Visualization of RNAs by Fluorescent In Situ Hybridization . . . . . . . . . . . . . . . . .
3
Thomas Dresselhaus and Andrea Bleckmann
2 Quantitative Fluorescence In Situ Hybridization Detection
of Plant mRNAs with Single-Molecule Resolution . . . . . . . . . . . . . . . . . . . . . . . . . . 23
Kun Huang, Mona Batish, Chong Teng, Alex Harkess,
Blake C. Meyers, and Jeffrey L. Caplan
3 Visualization of Endoplasmic Reticulum-Associated mRNA
in Mammalian Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 35
Jingze J. Wu and Alexander F. Palazzo
4 Simultaneous Detection of mRNA and Protein in S. cerevisiae
by Single-Molecule FISH and Immunofluorescence . . . . . . . . . . . . . . . . . . . . . . . . . 51
Evelina Tutucci and Robert H. Singer
PART II IN VIVO IMAGING OF RNA TRANSPORT AND LOCALIZATION
5 Development and Applications of Fluorogen/Light-Up RNA Aptamer
Pairs for RNA Detection and More . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 73
Michael Ryckelynck
6 Visualization of Transiently Expressed mRNA in Plants Using MS2 . . . . . . . . . . . 103
Eduardo Jose ´ Pen ˜a and Manfred Heinlein
7 New Generations of MS2 Variants and MCP Fusions to Detect
Single mRNAs in Living Eukaryotic Cells . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 121
Xavier Pichon, Marie-Ce ´cile Robert, Edouard Bertrand,
Robert H. Singer, and Evelina Tutucci
8 In Vivo Imaging of Mobile mRNAs in Plants Using MS2 . . . . . . . . . . . . . . . . . . . . 145
Kai-Ren Luo, Nien-Chen Huang, and Tien-Shin Yu
9 RNA Imaging with RNase-Inactivated Csy4 in Plants and Filamentous
Fungi. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 157
David Burnett, Alexander Lichius, and Jens Tilsner
PART III IMAGING AND ANALYSIS OF RNA UPTAKE AND TRANSPORT
BETWEEN CELLS
10 Utilizing Potato Virus X to Monitor RNA Movement. . . . . . . . . . . . . . . . . . . . . . . 181
Zhiming Yu, Sung Ki Cho, Pengcheng Zhang,
Yiguo Hong, and David J. Hannapel
vii