Single-cell isolation and gene analysis: Pitfalls and possibilities

International Journal of Molecular Sciences

Volumn 16, Issue 11, 2015, Pages 26832-26849

  Hodne, Kjetil ^a   Weltzien, Finn Arne ^a  

^a NORWEGIAN UNIVERSITY OF LIFE SCIENCES   (Norway)

Author keywords

Amplification qPCR; Analysis; Harvesting; RT; Single cell

Indexed keywords

3 [(3 CHOLAMIDOPROPYL)DIMETHYLAMMONIO] 1 PROPANESULFONIC ACID; DODECYL SULFATE SODIUM; TYLOXAPOL; MESSENGER RNA;

CAPILLARY ELECTROPHORESIS; CELL ISOLATION; DNA SYNTHESIS; ELECTROPHYSIOLOGY; FLUORESCENCE ACTIVATED CELL SORTING; GENETIC ANALYSIS; LASER MICRODISSECTION; MICROFLUIDICS; REAL TIME POLYMERASE CHAIN REACTION; REVERSE TRANSCRIPTION POLYMERASE CHAIN REACTION; REVIEW; RNA ISOLATION; SINGLE CELL ANALYSIS; SPECTROPHOTOMETRY; CELL SEPARATION; CHEMISTRY; DEVICES; ESCHERICHIA COLI; FLOW CYTOMETRY; GENE EXPRESSION PROFILING; GENE EXPRESSION REGULATION; GENETICS; HUMAN; LASER CAPTURE MICRODISSECTION; METABOLISM; PHENOTYPE; PROCEDURES; REVERSE TRANSCRIPTION; THERMUS;

CELL SEPARATION; ELECTROPHORESIS, CAPILLARY; ESCHERICHIA COLI; FLOW CYTOMETRY; GENE EXPRESSION PROFILING; GENE EXPRESSION REGULATION; HUMANS; LASER CAPTURE MICRODISSECTION; MICROFLUIDICS; PHENOTYPE; REAL-TIME POLYMERASE CHAIN REACTION; REVERSE TRANSCRIPTION; RNA, MESSENGER; SINGLE-CELL ANALYSIS; THERMUS;

EID: 84946897352     PISSN: 16616596     EISSN: 14220067     Source Type: Journal    
DOI: 10.3390/ijms161125996     Document Type: Review

References (116)

1. Ko, M.S.H.; Nakauchi, H.; Takahashi, N. The dose dependence of glucocorticoid-inducible gene-expression results from changes in the number of transcriptionally active templates. EMBO J. 1990, 9, 2835-2842. [PubMed]
2. Walters, M.C.; Fiering, S.; Eidemiller, J.; Magis, W.; Groudine, M.; Martin, D.I.K. Enhancers increase the probability but not the level of gene-expression. Proc. Natl. Acad. Sci. USA 1995, 92, 7125-7129. [CrossRef] [PubMed]
3. McAdams, H.H.; Arkin, A. Stochastic mechanisms in gene expression. Proc. Natl. Acad. Sci. USA 1997, 94, 814-819. [CrossRef] [PubMed]
4. Elowitz, M.B.; Levine, A.J.; Siggia, E.D.; Swain, P.S. Stochastic gene expression in a single cell. Science 2002, 297, 1183-1186. [CrossRef] [PubMed]
5. Norris, A.J.; Stirland, J.A.; McFerran, D.W.; Seymour, Z.C.; Spiller, D.G.; Loudon, A.S.I.; White, M.R.H.; Davis, J.R.E. Dynamic patterns of growth hormone gene transcription in individual living pituitary cells. Mol. Endocrinol. 2003, 17, 193-202. [CrossRef] [PubMed]
6. Bengtsson, M.; Stahlberg, A.; Rorsman, P.; Kubista, M. Gene expression profiling in single cells from the pancreatic islets of Langerhans reveals lognormal distribution of mRNA levels. Genome Res. 2005, 15, 1388-1392. [CrossRef] [PubMed]
7. Chubb, J.R.; Trcek, T.; Shenoy, S.M.; Singer, R.H. Transcriptional pulsing of a developmental gene. Curr. Biol. 2006, 16, 1018-1025. [CrossRef] [PubMed]
8. Raj, A.; Peskin, C.S.; Tranchina, D.; Vargas, D.Y.; Tyagi, S. Stochastic mRNA synthesis in mammalian cells. PLoS Biol. 2006, 4, 1707-1719. [CrossRef] [PubMed]
9. Raj, A.; van Oudenaarden, A. Nature, nurture, or chance: Stochastic gene expression and its consequences. Cell 2008, 135, 216-226. [CrossRef] [PubMed]
10. Method of the Year 2013. Methods to sequence the DNA and RNA of single cells are poised to transform many areas of biology and medicine. Nat. Methods 2014, 11, 1.
11. Malnic, B.; Hirono, J.; Sato, T.; Buck, L.B. Combinatorial receptor codes for odors. Cell 1999, 96, 713-723. [CrossRef]
12. Raghunathan, A.; Ferguson, H.R.; Bornarth, C.J.; Song, W.M.; Driscoll, M.; Lasken, R.S. Genomic DNA amplification from a single bacterium. Appl. Environ. Microbiol. 2005, 71, 3342-3347. [CrossRef] [PubMed]
13. Zhang, K.; Martiny, A.C.; Reppas, N.B.; Barry, K.W.; Malek, J.; Chisholm, S.W.; Church, G.M. Sequencing genomes from single cells by polymerase cloning. Nat. Biotechnol. 2006, 24, 680-686. [CrossRef] [PubMed]
14. Navin, N.; Kendall, J.; Troge, J.; Andrews, P.; Rodgers, L.; McIndoo, J.; Cook, K.; Stepansky, A.; Levy, D.; Esposito, D.; et al. Tumour evolution inferred by single-cell sequencing. Nature 2011, 472, 90-94. [CrossRef] [PubMed]
15. Zong, C.H.; Lu, S.J.; Chapman, A.R.; Xie, X.S. Genome-wide detection of single-nucleotide and copy-number variations of a single human cell. Science 2012, 338, 1622-1626. [CrossRef]
16. Kleppe, K.; Ohtsuka, E.; Kleppe, R.; Molineux, I.; Khorana, H.G. Studies on polynucleotides. XCVI. Repair replications of short synthetic DNA's as catalyzed by DNA polymerases. J. Mol. Biol. 1971, 56, 341-361. [CrossRef]
17. Rabinow, P. Making PCR: A Story of Biotechnology; University of Chicago Press: Chicago, IL, USA, 1996.
18. Saiki, R.K.; Scharf, S.; Faloona, F.; Mullis, K.B.; Horn, G.T.; Erlich, H.A.; Arnheim, N. Enzymatic amplification of β-globin genomic sequences and restriction site analysis for diagnosis of sickle-cell anemia. Science 1985, 230, 1350-1354. [CrossRef] [PubMed]
19. Mullis, K.; Faloona, F.; Scharf, S.; Saiki, R.; Horn, G.; Erlich, H. Specific enzymatic amplification of DNA in vitro-The polymerase chain-reaction. Cold Spring Harb. Symp. Quant. Biol. 1986, 51, 263-273. [CrossRef] [PubMed]
20. Saiki, R.K.; Gelfand, D.H.; Stoffel, S.; Scharf, S.J.; Higuchi, R.; Horn, G.T.; Mullis, K.B.; Erlich, H.A. Primer-directed enzymatic amplification of DNA with a thermostable DNA-polymerase. Science 1988, 239, 487-491. [CrossRef] [PubMed]
21. Chien, A.; Edgar, D.B.; Trela, J.M. Deoxyribonucleic-acid polymerase from extreme thermophile Thermus aquaticus. J. Bacteriol. 1976, 127, 1550-1557. [PubMed]
22. Brock, T.D.; Freeze, H. Thermus aquaticus gen. n. and sp. n., a nonsporulating extreme thermophile. J. Bacteriol. 1969, 98, 289-297. [PubMed]
23. Baltimore, D. RNA-dependent DNA polymerase in virions of RNA tumour viruses. Nature 1970, 226, 1209-1211. [CrossRef] [PubMed]
24. Temin, H.M.; Mizutani, S. Viral RNA-dependent DNA polymerase: RNA-dependent DNA polymerase in virions of rous sarcoma virus. Rev. Med. Virol. 1998, 8, 3-11. [CrossRef]
25. Verma, I.M.; Temple, G.F.; Baltimor, D.; Fan, H. In vitro synthesis of DNA complementary to rabbit reticulocyte 10S RNA. Nat. New Biol. 1972, 235, 163-167. [CrossRef] [PubMed]
26. Efstratiadis, A.; Kafatos, F.C.; Maxam, A.M.; Maniatis, T. Enzymatic in vitro synthesis of globin genes. Cell 1976, 7, 279-288. [CrossRef]
27. Fisher, M.P.; Dingman, C.W. Role of molecular conformation in determining electrophoretic properties of polynucleotides in agarose-acrylamide composite gels. Biochemistry 1971, 10, 1895-1899. [CrossRef] [PubMed]
28. Aaij, C.; Borst, P. The gel-electrophoresis of DNA. Biochim. Biophys. Acta 1972, 269, 192-200. [CrossRef]
29. Sharp, P.A.; Sugden, B.; Sambrook, J. Detection of two restriction endonuclease activities in haemophilus-parainfluenzae using analytical agarose-ethidium bromide electrophoresis. Biochemistry 1973, 12, 3055-3063. [CrossRef] [PubMed]
30. Li, H.; Gyllensten, U.B.; Cui, X.; Saiki, R.K.; Erlich, H.A.; Arnheim, N. Amplification and analysis of DNA sequences in single human sperm and diploid cells. Nature 1988, 335, 414-417. [CrossRef] [PubMed]
31. Brady, G.; Barbara, M.; Iscove, N.N. Representative in vitro cDNA amplification from individual hemopoietic cells and colonies. Methods Mol. Cell. Biol. 1990, 2, 17-25.
32. Vangelder, R.N.; Vonzastrow, M.E.; Yool, A.; Dement, W.C.; Barchas, J.D.; Eberwine, J.H. Amplified RNA synthesized from limited quantities of heterogeneous cDNA. Proc. Natl. Acad. Sci. USA 1990, 87, 1663-1667. [CrossRef]
33. Eberwine, J.; Yeh, H.; Miyashiro, K.; Cao, Y.X.; Nair, S.; Finnell, R.; Zettel, M.; Coleman, P. Analysis of gene-expression in single live neurons. Proc. Natl. Acad. Sci. USA 1992, 89, 3010-3014. [CrossRef] [PubMed]
34. Lambolez, B.; Audinat, E.; Bochet, P.; Crepel, F.; Rossier, J. AMPA receptor subunits expressed by single Purkinje cells. Neuron 1992, 9, 247-258. [CrossRef]
35. Higuchi, R.; Dollinger, G.;Walsh, P.S.; Griffith, R. Simultaneous amplification and detection of specific DNA sequences. Bio/Technology 1992, 10, 413-417. [CrossRef] [PubMed]
36. Higuchi, R.; Fockler, C.; Dollinger, G.; Watson, R. Kinetic PCR analysis: Real-time monitoring of DNA amplification reactions. Bio/Technology 1993, 11, 1026-1030. [CrossRef] [PubMed]
37. McPherson, M.J.; Hames, B.D.; Taylor, G.R. PCR 2: A Practical Approach; Oxford University Press: Oxford, UK, 1995.
38. Kainz, P. The PCR plateau phase-Towards an understanding of its limitations. Biochim. Biophys. Acta 2000, 1494, 23-27. [CrossRef]
39. Schmittgen, T.D.; Zakrajsek, B.A.; Mills, A.G.; Gorn, V.; Singer, M.J.; Reed, M.W. Quantitative reverse transcription-polymerase chain reaction to study mRNA decay: Comparison of endpoint and real-time methods. Anal. Biochem. 2000, 285, 194-204. [CrossRef] [PubMed]
40. t method. Methods 2001, 25, 402-408. [CrossRef] [PubMed]
41. Bustin, S.A. Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): Trends and problems. J. Mol. Endocrinol. 2002, 29, 17. [CrossRef]
42. Bustin, S.A.; Benes, V.; Garson, J.A.; Hellemans, J.; Huggett, J.; Kubista, M.; Mueller, R.; Nolan, T.; Pfaffl, M.W.; Shipley, G.L.; et al. The MIQE guidelines: Minimum information for publication of quantitative real-time PCR experiments. Clin. Chem. 2009, 55, 611-622. [CrossRef] [PubMed]
43. Karlsen, F.; Steen, H.B.; Nesland, J.M. SYBR green I DNA staining increases the detection sensitivity of viruses by polymerase chain-reaction. J. Virol. Methods 1995, 55, 153-156. [CrossRef]
44. Heid, C.A.; Stevens, J.; Livak, K.J.;Williams, P.M. Real time quantitative PCR. Genome Res. 1996, 6, 986-994. [CrossRef] [PubMed]
45. Wittwer, C.T.; Ririe, K.M.; Andrew, R.V.; David, D.A.; Gundry, R.A.; Balis, U.J. The LightCycler: A microvolume multisample fluorimeter with rapid temperature control. Biotechniques 1997, 22, 176-181. [PubMed]
46. Holland, P.M.; Abramson, R.D.; Watson, R.; Gelfand, D.H. Detection of specific polymerase chain-reaction product by utilizing the 51-31 exonuclease activity of Thermus aquaticus DNA-polymerase. Proc. Natl. Acad. Sci. USA 1991, 88, 7276-7280. [CrossRef] [PubMed]
47. Liu, P.; Mathies, R.A. Integrated microfluidic systems for high-performance genetic analysis. Trends Biotechnol. 2009, 27, 572-581. [CrossRef] [PubMed]
48. Nagarajan, N.; Pop, M. Sequence assembly demystified. Nat. Rev. Genet. 2013, 14, 157-167. [CrossRef] [PubMed]
49. Shapiro, E.; Biezuner, T.; Linnarsson, S. Single-cell sequencing-based technologies will revolutionize whole-organism science. Nat. Rev. Genet. 2013, 14, 618-630. [CrossRef] [PubMed]
50. Van Loo, P.; Voet, T. Single cell analysis of cancer genomes. Curr. Opin. Genet. Dev. 2014, 24, 82-91. [CrossRef] [PubMed]
51. Achim, K.; Pettit, J.-B.; Saraiva, L.R.; Gavriouchkina, D.; Larsson, T.; Arendt, D.; Marioni, J.C. High-throughput spatial mapping of single-cell RNA-seq data to tissue of origin. Nat. Biotechnol. 2015, 33, 503-509. [CrossRef] [PubMed]
52. Buettner, F.; Natarajan, K.N.; Casale, F.P.; Proserpio, V.; Scialdone, A.; Theis, F.J.; Teichmann, S.A.; Marioni, J.C.; Stegie, O. Computational analysis of cell-to-cell heterogeneity in single-cell RNA-sequencing data reveals hidden subpopulations of cells. Nat. Biotechnol. 2015, 33, 155-160. [CrossRef] [PubMed]
53. Crosetto, N.; Bienko, M.; van Oudenaarden, A. Spatially resolved transcriptomics and beyond. Nat. Rev. Genet. 2015, 16, 57-66. [CrossRef] [PubMed]
54. Darmanis, S.; Sloan, S.A.; Zhang, Y.; Enge, M.; Caneda, C.; Shuer, L.M.; Gephart, M.G.H.; Barres, B.A.; Quake, S.R. A survey of human brain transcriptome diversity at the single cell level. Proc. Natl. Acad. Sci. USA 2015, 112, 7285-7290. [CrossRef] [PubMed]
55. Stegle, O.; Teichmann, S.A.; Marioni, J.C. Computational and analytical challenges in single-cell transcriptomics. Nat. Rev. Genet. 2015, 16, 133-145. [CrossRef] [PubMed]
56. EmmertBuck, M.R.; Bonner, R.F.; Smith, P.D.; Chuaqui, R.F.; Zhuang, Z.P.; Goldstein, S.R.; Weiss, R.A.; Liotta, L.A. Laser capture microdissection. Science 1996, 274, 998-1001. [CrossRef]
57. Bonner, R.F.; EmmertBuck, M.; Cole, K.; Pohida, T.; Chuaqui, R.; Goldstein, S.; Liotta, L.A. Laser capture microdissection: Molecular analysis of tissue. Science 1997, 278, 1481-1483. [CrossRef] [PubMed]
58. Schutze, K.; Lahr, G. Identification of expressed genes by laser-mediated manipulation of single cells. Nat. Biotechnol. 1998, 16, 737-742. [CrossRef] [PubMed]
59. Podgorny, O.V. Live cell isolation by laser microdissection with gravity transfer. J. Biomed. Opt. 2013, 18, 8. [CrossRef] [PubMed]
60. Herzenberg, L.A.; Sweet, R.G. Fluorescence-activated cell sorting. Sci. Am. 1976, 234, 108-117. [CrossRef] [PubMed]
61. Herzenberg, L.A.; Parks, D.; Sahaf, B.; Perez, O.; Roederer, M. The history and future of the fluorescence activated cell sorter and flow cytometry: A view from Stanford. Clin. Chem. 2002, 48, 1819-1827. [PubMed]
62. Wheeler, A.R.; Throndset, W.R.; Whelan, R.J.; Leach, A.M.; Zare, R.N.; Liao, Y.H.; Farrell, K.; Manger, I.D.; Daridon, A. Microfluidic device for single-cell analysis. Anal. Chem. 2003, 75, 3581-3586. [CrossRef] [PubMed]
63. Ottesen, E.A.; Hong, J.W.; Quake, S.R.; Leadbetter, J.R. Microfluidic digital PCR enables multigene analysis of individual environmental bacteria. Science 2006, 314, 1464-1467. [CrossRef] [PubMed]
64. Warren, L.; Bryder, D.; Weissman, I.L.; Quake, S.R. Transcription factor profiling in individual hematopoietic progenitors by digital RT-PCR. Proc. Natl. Acad. Sci. USA 2006, 103, 17807-17812. [CrossRef] [PubMed]
65. Whitesides, G.M. The origins and the future of microfluidics. Nature 2006, 442, 368-373. [CrossRef] [PubMed]
66. Wu, A.R.; Neff, N.F.; Kalisky, T.; Dalerba, P.; Treutlein, B.; Rothenberg, M.E.; Mburu, F.M.; Mantalas, G.L.; Sim, S.; Clarke, M.F.; Quake, S.R. Quantitative assessment of single-cell RNA-sequencing methods. Nat. Methods 2014, 11, 41-46. [CrossRef] [PubMed]
67. Unger, M.A.; Chou, H.P.; Thorsen, T.; Scherer, A.; Quake, S.R. Monolithic microfabricated valves and pumps by multilayer soft lithography. Science 2000, 288, 113-116. [CrossRef] [PubMed]
68. Joensson, H.N.; Svahn, H.A. Droplet microfluidics-A tool for single-cell analysis. Angew. Chem. Int. Ed. 2012, 51, 12176-12192. [CrossRef] [PubMed]
69. Seemann, R.; Brinkmann, M.; Pfohl, T.; Herminghaus, S. Droplet based microfluidics. Rep. Prog. Phys. 2012, 75, 016601. [CrossRef] [PubMed]
70. Pantoja, R.; Nagarah, J.M.; Starace, D.M.; Melosh, N.A.; Blunck, R.; Bezanilla, F.; Heath, J.R. Silicon chip-based patch-clamp electrodes integrated with PDMS microfluidics. Biosens. Bioelectron. 2004, 20, 509-517. [CrossRef] [PubMed]
71. Hong, J.W.; Studer, V.; Hang, G.; Anderson, W.F.; Quake, S.R. A nanoliter-scale nucleic acid processor with parallel architecture. Nat. Biotechnol. 2004, 22, 435-439. [CrossRef] [PubMed]
72. Wu, H.K.; Wheeler, A.; Zare, R.N. Chemical cytometry on a picoliter-scale integrated microfluidic chip. Proc. Natl. Acad. Sci. USA 2004, 101, 12809-12813. [CrossRef] [PubMed]
73. Walch, A.; Specht, K.; Smida, J.; Aubele, M.; Zitzelsberger, H.; Hofler, H.;Werner, M. Tissue microdissection techniques in quantitative genome and gene expression analyses. Histochem. Cell Biol. 2001, 115, 269-276. [PubMed]
74. Guo, G.J.; Huss, M.; Tong, G.Q.; Wang, C.Y.; Sun, L.L.; Clarke, N.D.; Robson, P. Resolution of Cell Fate Decisions Revealed by Single-Cell Gene Expression Analysis from Zygote to Blastocyst. Dev. Cell 2010, 18, 675-685. [CrossRef] [PubMed]
75. Hodne, K.; Haug, T.M.;Weltzien, F.A. Single-cell qPCR on dispersed primary pituitary cells-An optimized protocol. BMC Mol. Biol. 2010, 11, 82. [CrossRef] [PubMed]
76. Citri, A.; Pang, Z.P.P.; Sudhof, T.C.; Wernig, M.; Malenka, R.C. Comprehensive qPCR profiling of gene expression in single neuronal cells. Nat. Protoc. 2012, 7, 118-127. [CrossRef] [PubMed]
77. Brown, R.B.; Audet, J. Current techniques for single-cell lysis. J. R. Soc. Interface 2008, 5, S131-S138. [CrossRef] [PubMed]
78. Svec, D.; Andersson, D.; Pekny, M.; Sjoback, R.; Kubista, M.; Stahlberg, A. Direct cell lysis for single-cell gene expression profiling. Front. Oncol. 2013, 3, 274. [CrossRef] [PubMed]
79. Stahlberg, A.; Kubista, M. The workflow of single-cell expression profiling using quantitative real-time PCR. Expert Rev. Mol. Diagn. 2014, 14, 323-331. [CrossRef] [PubMed]
80. Vonhippel, P.H.; Wong, K.Y. Neutral salts-Generality of their effects on stability of macromolecular conformations. Science 1964, 145, 577-580. [CrossRef] [PubMed]
81. Nozaki, Y.; Tanford, C. Solubility of amino acids, diglycine, and triglycine in aqueous guanidine hydrochloride solutions. J. Biol. Chem. 1970, 245, 1648-1652. [PubMed]
82. Gordon, J.A. Denaturation of globular proteins. Interaction of guanidinium salts with three proteins. Biochemistry 1972, 11, 1862-1870. [CrossRef] [PubMed]
83. Bengtsson, M.; Hemberg, M.; Rorsman, P.; Stahlberg, A. Quantification of mRNA in single cells and modelling of RT-qPCR induced noise. BMC Mol. Biol. 2008, 9, 11. [CrossRef] [PubMed]
84. Geselowitz, D.A.; Neckers, L.M. Bovine serum-albumin is a major oligonucleotide-binding protein found on the surface of cultured-cells. Antisense Res. Dev. 1995, 5, 213-217. [PubMed]
85. Kreader, C.A. Relief of amplification inhibition in PCR with bovine serum albumin or T4 gene 32 protein. Appl. Environ. Microbiol. 1996, 62, 1102-1106. [PubMed]
86. Wilson, I.G. Inhibition and facilitation of nucleic acid amplification. Appl. Environ. Microbiol. 1997, 63, 3741-3751. [PubMed]
87. Abu Al-Soud, W.; Radstrom, P. Effects of amplification facilitators on diagnostic PCR in the presence of blood, feces, and meat. J. Clin. Microbiol. 2000, 38, 4463-4470. [PubMed]
88. Arnedo, A.; Espuelas, S.; Irache, J.M. Albumin nanoparticles as carriers for a phosphodiester oligonucleotide. Int. J. Pharm. 2002, 244, 59-72. [CrossRef]
89. Farell, E.M.; Alexandre, G. Bovine serum albumin further enhances the effects of organic solvents on increased yield of polymerase chain reaction of GC-rich templates. BMC Res. Notes 2012, 5, 257. [CrossRef] [PubMed]
90. Deprez, R.H.L.; Fijnvandraat, A.C.; Ruijter, J.M.; Moorman, A.F.M. Sensitivity and accuracy of quantitative real-time polymerase chain reaction using SYBR green I depends on cDNA synthesis conditions. Anal. Biochem. 2002, 307, 63-69. [CrossRef]
91. Stahlberg, A.; Hakansson, J.; Xian, X.J.; Semb, H.; Kubista, M. Properties of the reverse transcription reaction in mRNA quantification. Clin. Chem. 2004, 50, 509-515. [CrossRef] [PubMed]
92. Islam, S.; Zeisel, A.; Joost, S.; la Manno, G.; Zajac, P.; Kasper, M.; Lonnerberg, P.; Linnarsson, S. Quantitative single-cell RNA-seq with unique molecular identifiers. Nat. Methods 2014, 11, 163-166. [CrossRef] [PubMed]
93. Ikeda, R.A. The efficiency of promoter clearance distinguishes T7 class-II and class-III promoters. J. Biol. Chem. 1992, 267, 11322-11328. [PubMed]
94. Ikeda, R.A.; Lin, A.C.; Clarke, J. Initiation of transcription by T7-RNA polymerase at its natural promoters. J. Biol. Chem. 1992, 267, 2640-2649.
95. Pabon, C.; Modrusan, Z.; Ruvolo, M.V.; Coleman, I.M.; Daniel, S.; Yue, H.; Arnold, L.J.; Reynolds, M.A. Optimized T7 amplification system for microarray analysis. Biotechniques 2001, 31, 874-879. [PubMed]
96. Wang, J.; Hu, L.; Hamilton, S.R.; Coombes, K.R.; Zhang, W. RNA amplification strategies for cDNA microarray experiments. BioTechniques 2003, 34, 394-400. [PubMed]
97. Moll, P.R.; Duschl, J.; Richter, K. Optimized RNA amplification using T7-RNA-polymerase based in vitro transcription. Anal. Biochem. 2004, 334, 164-174. [CrossRef] [PubMed]
98. Brady, G.; Billia, F.; Knox, J.; Hoang, T.; Kirsch, I.R.; Voura, E.B.; Hawley, R.G.; Cumming, R.; Buchwald, M.; Siminovitch, K.; et al. analysis of gene-expression in a complex differentiation hierarchy by global amplification of cDNA from single cells. Curr. Biol. 1995, 5, 909-922. [CrossRef]
99. Iscove, N.N.; Barbara, M.; Gu, M.; Gibson, M.; Modi, C.; Winegarden, N. Representation is faithfully preserved in global cDNA amplified exponentially from sub-picogram quantities of mRNA. Nat. Biotechnol. 2002, 20, 940-943. [CrossRef] [PubMed]
100. Subkhankulova, T.; Livesey, F.J. Comparative evaluation of linear and exponential amplification techniques for expression profiling at the single-cell level. Genome Biol. 2006, 7, 16. [CrossRef] [PubMed]
101. Lang, J.E.; Magbanua, M.J.M.; Scott, J.H.; Makrigiorgos, G.M.; Wang, G.; Federman, S.; Esserman, L.J.; Park, J.W.; Haqq, C.M. A comparison of RNA amplification techniques at sub-nanogram input concentration. Bmc Genom. 2009, 10, 12. [CrossRef] [PubMed]
102. Zhu, Y.Y.; Machleder, E.M.; Chenchik, A.; Li, R.; Siebert, P.D. Reverse transcriptase template switching: A SMART™ approach for full-length cDNA library construction. Biotechniques 2001, 30, 892-897. [PubMed]
103. Tang, F.C.; Barbacioru, C.; Wang, Y.Z.; Nordman, E.; Lee, C.; Xu, N.L.; Wang, X.H.; Bodeau, J.; Tuch, B.B.; Siddiqui, A.; et al. mRNA-Seq whole-transcriptome analysis of a single cell. Nat. Methods 2009, 6, 377-382. [CrossRef] [PubMed]
104. Islam, S.; Kjallquist, U.; Moliner, A.; Zajac, P.; Fan, J.B.; Lonnerberg, P.; Linnarsson, S. Characterization of the single-cell transcriptional landscape by highly multiplex RNA-seq. Genome Res. 2011, 21, 1160-1167. [CrossRef] [PubMed]
105. Hashimshony, T.; Wagner, F.; Sher, N.; Yanai, I. CEL-Seq: Single-cell RNA-seq by multiplexed linear amplification. Cell Rep. 2012, 2, 666-673. [CrossRef] [PubMed]
106. Ramskold, D.; Luo, S.J.; Wang, Y.C.; Li, R.; Deng, Q.L.; Faridani, O.R.; Daniels, G.A.; Khrebtukova, I.; Loring, J.F.; Laurent, L.C.; et al. Full-length mRNA-Seq from single-cell levels of RNA and individual circulating tumor cells. Nat. Biotechnol. 2012, 30, 777-782. [CrossRef] [PubMed]
107. Sasagawa, Y.; Nikaido, I.; Hayashi, T.; Danno, H.; Uno, K.D.; Imai, T.; Ueda, H.R. Quartz-Seq: A highly reproducible and sensitive single-cell RNA sequencing method, reveals non-genetic gene-expression heterogeneity. Genome Biol. 2013, 14, 17. [CrossRef] [PubMed]
108. Koch, W.H. Technology platforms for pharmacogenomic diagnostic assays. Nat. Rev. Drug Discov. 2004, 3, 749-761. [CrossRef] [PubMed]
109. Persson, K.; Hamby, K.; Ugozzoli, L.A. Four-color multiplex reverse transcription polymerase chain reaction-Overcoming its limitations. Anal. Biochem. 2005, 344, 33-42. [CrossRef] [PubMed]
110. Primer3plus. Available online: http://www.bioinformatics.nl/cgi-bin/primer3plus/primer3plus.cgi (accessed on 5 November 2015).
111. Vector NTI. Available online: http://www.lifetechnologies.com/no/en/home/life-science/cloning/ vector-nti-software.html (accessed on 5 November 2015).
112. Chandler, D.P.;Wagnon, C.A.; Bolton, H. Reverse transcriptase (RT) inhibition of PCR at low concentrations of template and its implications for quantitative RT-PCR. Appl. Environ. Microbiol. 1998, 64, 669-677. [PubMed]
113. Liss, B. Improved quantitative real-time RT-PCR for expression profiling of individual cells. Nucleic Acids Res. 2002, 30, 9. [CrossRef]
114. Nolan, T.; Hands, R.E.; Ogunkolade, W.; Bustin, S.A. SPUD: A quantitative PCR assay for the detection of inhibitors in nucleic acid preparations. Anal. Biochem. 2006, 351, 308-310. [CrossRef] [PubMed]
115. Singhal, R.; Orynbayeva, Z.; Sundaram, R.V.K.; Niu, J.J.; Bhattacharyya, S.; Vitol, E.A.; Schrlau, M.G.; Papazoglou, E.S.; Friedman, G.; Gogotsi, Y. Multifunctional carbon-nanotube cellular endoscopes. Nat. Nanotechnol. 2011, 6, 57-64. [CrossRef] [PubMed]
116. Schrlau, M.G.; Dun, N.J.; Bau, H.H. Cell electrophysiology with carbon nanopipettes. ACS Nano 2009, 3, 563-568. [CrossRef] [PubMed]