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High-Efficiency Cloning of Full-Length cDNA
Open accessAbstract
A widely recognized difficulty of presently used methods for cDNA cloning is obtaining cDNA segments that contain the entire nucleotide sequence of the corresponding mRNA. The cloning procedure described here mitigates this shortcoming. Of the 10(5) plasmid-cDNA recombinants obtained per microgram of rabbit reticulocyte mRNA, about 10% contained a complete alpha- of beta-globin mRNA sequence, and at least 30 to 50%, but very likely more, contained the entire globin coding regions. We attribute the high efficiency of cloning full- or nearly full-length cDNA to (i) the fact that the plasmid DNA vector itself serves as the primer for first- and second-strand cDNA synthesis, (ii) the lack of any nuclease treatment of the products, and (iii) the fact that one of the steps in the procedure results in preferential cloning of recombinants with full-length cDNA's over those with truncated cDNA's.
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Citations
Cited by
The human LDL receptor: A cysteine-rich protein with multiple Alu sequences in its mRNA
Yamamoto, T., Davis, C. G., Brown, M. S., Schneider, W. J., Casey, M. L., Goldstein, J. L., & Russell, D. W. (1984) · Cell · doi:10.1016/0092-8674(84)90188-0 · Cited by 1,382
Cloning and characterization of the cDNA encoding a novel human pre-B-cell colony-enhancing factor.
Samal, B., Sun, Y., Stearns, G., Xie, C., Suggs, S., & McNiece, I. (1994) · Molecular and Cellular Biology · doi:10.1128/mcb.14.2.1431 · Cited by 1,019
Easy identification of cDNA clones.
Rüther, U., & Müller-Hill, B. (1983) · The EMBO Journal · doi:10.1002/j.1460-2075.1983.tb01659.x · Cited by 573
Takahashi, S., Kawarabayasi, Y., Nakai, T., Sakai, J., & Yamamoto, T. (1992) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.89.19.9252 · Cited by 549
Large scale cDNA sequencing for analysis of quantitative and qualitative aspects of gene expression
Okubo, K., Hori, N., Matoba, R., Niiyama, T., Fukushima, A., Kojima, Y., & Matsubara, K. (1992) · Nature Genetics · doi:10.1038/ng1192-173 · Cited by 522
Metadata
- Authors
- Okayama, H., & Berg, P.
- Venue
- Molecular and Cellular Biology
- Publisher
- Informa UK Limited
- Published
- 1982-02-01
- Volume
- 2
- Issue
- 2
- Pages
- 161-170
- ISSN
- 0270-7306 (ISSN-L)
- 1098-5549
- OpenAlex
- W2038346712
- Cited by
- 1,267
- References
- 0
- FWCI
- 38.46
- Type
- journal article
- Language
- en
- Open access
- green
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