Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1997 Jul;17(7):3786–3798. doi: 10.1128/mcb.17.7.3786

Purification and characterization of FBI-1, a cellular factor that binds to the human immunodeficiency virus type 1 inducer of short transcripts.

F Pessler 1, P S Pendergrast 1, N Hernandez 1
PMCID: PMC232230  PMID: 9199312

Abstract

The human immunodeficiency virus (HIV-1) promoter directs the synthesis of two classes of RNA molecules, short transcripts and full-length transcripts. The synthesis of short transcripts depends on a bipartite DNA element, the inducer of short transcripts (IST), located in large part downstream of the HIV-1 start site of transcription. IST does not require any viral product for function and is thought to direct the assembly of transcription complexes that are incapable of efficient elongation. Nothing is known, however, about the biochemical mechanisms that mediate IST function. Here, we report the identification and purification of a factor that binds specifically to the IST. This factor, FBI-1, recognizes a large bipartite binding site that coincides with the bipartite IST element. It is constituted at least in part by an 86-kDa polypeptide that can be specifically cross-linked to IST. FBI-1 also binds to promoter and attenuation regions of a number of cellular and viral transcription units that are regulated by a transcription elongation block. This observation, together with the observation that the binding of FBI-1 to IST mutants correlates with the ability of these mutants to direct IST function, suggests that FBI-1 may be involved in the establishment of abortive transcription complexes.

Full Text

The Full Text of this article is available as a PDF (2.4 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ares M., Jr, Mangin M., Weiner A. M. Orientation-dependent transcriptional activator upstream of a human U2 snRNA gene. Mol Cell Biol. 1985 Jul;5(7):1560–1570. doi: 10.1128/mcb.5.7.1560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Ashfield R., Patel A. J., Bossone S. A., Brown H., Campbell R. D., Marcu K. B., Proudfoot N. J. MAZ-dependent termination between closely spaced human complement genes. EMBO J. 1994 Dec 1;13(23):5656–5667. doi: 10.1002/j.1460-2075.1994.tb06904.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bender T. P., Thompson C. B., Kuehl W. M. Differential expression of c-myb mRNA in murine B lymphomas by a block to transcription elongation. Science. 1987 Sep 18;237(4821):1473–1476. doi: 10.1126/science.3498214. [DOI] [PubMed] [Google Scholar]
  4. Bentley D. L., Groudine M. A block to elongation is largely responsible for decreased transcription of c-myc in differentiated HL60 cells. Nature. 1986 Jun 12;321(6071):702–706. doi: 10.1038/321702a0. [DOI] [PubMed] [Google Scholar]
  5. Bentley D. L., Groudine M. Sequence requirements for premature termination of transcription in the human c-myc gene. Cell. 1988 Apr 22;53(2):245–256. doi: 10.1016/0092-8674(88)90386-8. [DOI] [PubMed] [Google Scholar]
  6. Bossone S. A., Asselin C., Patel A. J., Marcu K. B. MAZ, a zinc finger protein, binds to c-MYC and C2 gene sequences regulating transcriptional initiation and termination. Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7452–7456. doi: 10.1073/pnas.89.16.7452. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Brown S. A., Imbalzano A. N., Kingston R. E. Activator-dependent regulation of transcriptional pausing on nucleosomal templates. Genes Dev. 1996 Jun 15;10(12):1479–1490. doi: 10.1101/gad.10.12.1479. [DOI] [PubMed] [Google Scholar]
  8. Buonaguro L., Buonaguro F. M., Giraldo G., Ensoli B. The human immunodeficiency virus type 1 Tat protein transactivates tumor necrosis factor beta gene expression through a TAR-like structure. J Virol. 1994 Apr;68(4):2677–2682. doi: 10.1128/jvi.68.4.2677-2682.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Carvalho M., Kirkland M., Derse D. Protein interactions with DNA elements in variant equine infectious anemia virus enhancers and their impact on transcriptional activity. J Virol. 1993 Nov;67(11):6586–6595. doi: 10.1128/jvi.67.11.6586-6595.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chinsky J. M., Maa M. C., Ramamurthy V., Kellems R. E. Adenosine deaminase gene expression. Tissue-dependent regulation of transcriptional elongation. J Biol Chem. 1989 Aug 25;264(24):14561–14565. [PubMed] [Google Scholar]
  11. Eick D., Berger R., Polack A., Bornkamm G. W. Transcription of c-myc in human mononuclear cells is regulated by an elongation block. Oncogene. 1987;2(1):61–65. [PubMed] [Google Scholar]
  12. Eick D., Bornkamm G. W. Transcriptional arrest within the first exon is a fast control mechanism in c-myc gene expression. Nucleic Acids Res. 1986 Nov 11;14(21):8331–8346. doi: 10.1093/nar/14.21.8331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Feinberg M. B., Baltimore D., Frankel A. D. The role of Tat in the human immunodeficiency virus life cycle indicates a primary effect on transcriptional elongation. Proc Natl Acad Sci U S A. 1991 May 1;88(9):4045–4049. doi: 10.1073/pnas.88.9.4045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fort P., Rech J., Vie A., Piechaczyk M., Bonnieu A., Jeanteur P., Blanchard J. M. Regulation of c-fos gene expression in hamster fibroblasts: initiation and elongation of transcription and mRNA degradation. Nucleic Acids Res. 1987 Jul 24;15(14):5657–5667. doi: 10.1093/nar/15.14.5657. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fried M., Crothers D. M. Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res. 1981 Dec 11;9(23):6505–6525. doi: 10.1093/nar/9.23.6505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Haley J. D., Waterfield M. D. Contributory effects of de novo transcription and premature transcript termination in the regulation of human epidermal growth factor receptor proto-oncogene RNA synthesis. J Biol Chem. 1991 Jan 25;266(3):1746–1753. [PubMed] [Google Scholar]
  17. Hay N., Skolnik-David H., Aloni Y. Attenuation in the control of SV40 gene expression. Cell. 1982 May;29(1):183–193. doi: 10.1016/0092-8674(82)90102-7. [DOI] [PubMed] [Google Scholar]
  18. Henry R. W., Sadowski C. L., Kobayashi R., Hernandez N. A TBP-TAF complex required for transcription of human snRNA genes by RNA polymerase II and III. Nature. 1995 Apr 13;374(6523):653–656. doi: 10.1038/374653a0. [DOI] [PubMed] [Google Scholar]
  19. Huang C. J., Goodart S. A., Rice M. K., Guzowski J. F., Wagner E. K. Mutational analysis of sequences downstream of the TATA box of the herpes simplex virus type 1 major capsid protein (VP5/UL19) promoter. J Virol. 1993 Sep;67(9):5109–5116. doi: 10.1128/jvi.67.9.5109-5116.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Jeffers M., Pellicer A. Multiple intragenic elements regulate the expression of the murine N-ras gene. Oncogene. 1992 Nov;7(11):2115–2123. [PubMed] [Google Scholar]
  21. Jones K. A. HIV trans-activation and transcription control mechanisms. New Biol. 1989 Nov;1(2):127–135. [PubMed] [Google Scholar]
  22. Jones K. A., Luciw P. A., Duchange N. Structural arrangements of transcription control domains within the 5'-untranslated leader regions of the HIV-1 and HIV-2 promoters. Genes Dev. 1988 Sep;2(9):1101–1114. doi: 10.1101/gad.2.9.1101. [DOI] [PubMed] [Google Scholar]
  23. Jones K. A., Peterlin B. M. Control of RNA initiation and elongation at the HIV-1 promoter. Annu Rev Biochem. 1994;63:717–743. doi: 10.1146/annurev.bi.63.070194.003441. [DOI] [PubMed] [Google Scholar]
  24. Kao S. Y., Calman A. F., Luciw P. A., Peterlin B. M. Anti-termination of transcription within the long terminal repeat of HIV-1 by tat gene product. Nature. 1987 Dec 3;330(6147):489–493. doi: 10.1038/330489a0. [DOI] [PubMed] [Google Scholar]
  25. Kennedy G. C., Rutter W. J. Pur-1, a zinc-finger protein that binds to purine-rich sequences, transactivates an insulin promoter in heterologous cells. Proc Natl Acad Sci U S A. 1992 Dec 1;89(23):11498–11502. doi: 10.1073/pnas.89.23.11498. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Krumm A., Hickey L. B., Groudine M. Promoter-proximal pausing of RNA polymerase II defines a general rate-limiting step after transcription initiation. Genes Dev. 1995 Mar 1;9(5):559–572. doi: 10.1101/gad.9.5.559. [DOI] [PubMed] [Google Scholar]
  27. Krumm A., Meulia T., Brunvand M., Groudine M. The block to transcriptional elongation within the human c-myc gene is determined in the promoter-proximal region. Genes Dev. 1992 Nov;6(11):2201–2213. doi: 10.1101/gad.6.11.2201. [DOI] [PubMed] [Google Scholar]
  28. Krumm A., Meulia T., Groudine M. Common mechanisms for the control of eukaryotic transcriptional elongation. Bioessays. 1993 Oct;15(10):659–665. doi: 10.1002/bies.950151005. [DOI] [PubMed] [Google Scholar]
  29. Kunkel G. R., Pederson T. Upstream elements required for efficient transcription of a human U6 RNA gene resemble those of U1 and U2 genes even though a different polymerase is used. Genes Dev. 1988 Feb;2(2):196–204. doi: 10.1101/gad.2.2.196. [DOI] [PubMed] [Google Scholar]
  30. Kyöstiö S. R., Wonderling R. S., Owens R. A. Negative regulation of the adeno-associated virus (AAV) P5 promoter involves both the P5 rep binding site and the consensus ATP-binding motif of the AAV Rep68 protein. J Virol. 1995 Nov;69(11):6787–6796. doi: 10.1128/jvi.69.11.6787-6796.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Laspia M. F., Rice A. P., Mathews M. B. HIV-1 Tat protein increases transcriptional initiation and stabilizes elongation. Cell. 1989 Oct 20;59(2):283–292. doi: 10.1016/0092-8674(89)90290-0. [DOI] [PubMed] [Google Scholar]
  32. Laspia M. F., Rice A. P., Mathews M. B. Synergy between HIV-1 Tat and adenovirus E1A is principally due to stabilization of transcriptional elongation. Genes Dev. 1990 Dec;4(12B):2397–2408. doi: 10.1101/gad.4.12b.2397. [DOI] [PubMed] [Google Scholar]
  33. Laspia M. F., Wendel P., Mathews M. B. HIV-1 Tat overcomes inefficient transcriptional elongation in vitro. J Mol Biol. 1993 Aug 5;232(3):732–746. doi: 10.1006/jmbi.1993.1427. [DOI] [PubMed] [Google Scholar]
  34. Lee H., Kraus K. W., Wolfner M. F., Lis J. T. DNA sequence requirements for generating paused polymerase at the start of hsp70. Genes Dev. 1992 Feb;6(2):284–295. doi: 10.1101/gad.6.2.284. [DOI] [PubMed] [Google Scholar]
  35. Lee K. J., Lee A. H., Sung Y. C. Multiple positive and negative cis-acting elements that mediate transactivation by bel1 in the long terminal repeat of human foamy virus. J Virol. 1993 Apr;67(4):2317–2326. doi: 10.1128/jvi.67.4.2317-2326.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Lobo S. M., Lister J., Sullivan M. L., Hernandez N. The cloned RNA polymerase II transcription factor IID selects RNA polymerase III to transcribe the human U6 gene in vitro. Genes Dev. 1991 Aug;5(8):1477–1489. doi: 10.1101/gad.5.8.1477. [DOI] [PubMed] [Google Scholar]
  37. Löchelt M., Muranyi W., Flügel R. M. Human foamy virus genome possesses an internal, Bel-1-dependent and functional promoter. Proc Natl Acad Sci U S A. 1993 Aug 1;90(15):7317–7321. doi: 10.1073/pnas.90.15.7317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Maa M. C., Chinsky J. M., Ramamurthy V., Martin B. D., Kellems R. E. Identification of transcription stop sites at the 5' and 3' ends of the murine adenosine deaminase gene. J Biol Chem. 1990 Jul 25;265(21):12513–12519. [PubMed] [Google Scholar]
  39. Maderious A., Chen-Kiang S. Pausing and premature termination of human RNA polymerase II during transcription of adenovirus in vivo and in vitro. Proc Natl Acad Sci U S A. 1984 Oct;81(19):5931–5935. doi: 10.1073/pnas.81.19.5931. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Marciniak R. A., Calnan B. J., Frankel A. D., Sharp P. A. HIV-1 Tat protein trans-activates transcription in vitro. Cell. 1990 Nov 16;63(4):791–802. doi: 10.1016/0092-8674(90)90145-5. [DOI] [PubMed] [Google Scholar]
  41. Marciniak R. A., Sharp P. A. HIV-1 Tat protein promotes formation of more-processive elongation complexes. EMBO J. 1991 Dec;10(13):4189–4196. doi: 10.1002/j.1460-2075.1991.tb04997.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Margolis D. M., Somasundaran M., Green M. R. Human transcription factor YY1 represses human immunodeficiency virus type 1 transcription and virion production. J Virol. 1994 Feb;68(2):905–910. doi: 10.1128/jvi.68.2.905-910.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Marthas M. L., Ramos R. A., Lohman B. L., Van Rompay K. K., Unger R. E., Miller C. J., Banapour B., Pedersen N. C., Luciw P. A. Viral determinants of simian immunodeficiency virus (SIV) virulence in rhesus macaques assessed by using attenuated and pathogenic molecular clones of SIVmac. J Virol. 1993 Oct;67(10):6047–6055. doi: 10.1128/jvi.67.10.6047-6055.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Means A. L., Farnham P. J. Transcription initiation from the dihydrofolate reductase promoter is positioned by HIP1 binding at the initiation site. Mol Cell Biol. 1990 Feb;10(2):653–661. doi: 10.1128/mcb.10.2.653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Mechti N., Piechaczyk M., Blanchard J. M., Jeanteur P., Lebleu B. Sequence requirements for premature transcription arrest within the first intron of the mouse c-fos gene. Mol Cell Biol. 1991 May;11(5):2832–2841. doi: 10.1128/mcb.11.5.2832. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Meulia T., Krumm A., Spencer C., Groudine M. Sequences in the human c-myc P2 promoter affect the elongation and premature termination of transcripts initiated from the upstream P1 promoter. Mol Cell Biol. 1992 Oct;12(10):4590–4600. doi: 10.1128/mcb.12.10.4590. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Miller H., Asselin C., Dufort D., Yang J. Q., Gupta K., Marcu K. B., Nepveu A. A cis-acting element in the promoter region of the murine c-myc gene is necessary for transcriptional block. Mol Cell Biol. 1989 Dec;9(12):5340–5349. doi: 10.1128/mcb.9.12.5340. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Mok M., Maderious A., Chen-Kiang S. Premature termination by human RNA polymerase II occurs temporally in the adenovirus major late transcriptional unit. Mol Cell Biol. 1984 Oct;4(10):2031–2040. doi: 10.1128/mcb.4.10.2031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Narayan S., Widen S. G., Beard W. A., Wilson S. H. RNA polymerase II transcription. Rate of promoter clearance is enhanced by a purified activating transcription factor/cAMP response element-binding protein. J Biol Chem. 1994 Apr 29;269(17):12755–12763. [PubMed] [Google Scholar]
  50. Nyborg J. K., Dynan W. S., Chen I. S., Wachsman W. Binding of host-cell factors to DNA sequences in the long terminal repeat of human T-cell leukemia virus type I: implications for viral gene expression. Proc Natl Acad Sci U S A. 1988 Mar;85(5):1457–1461. doi: 10.1073/pnas.85.5.1457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Pendergrast P. S., Hernandez N. RNA-targeted activators, but not DNA-targeted activators, repress the synthesis of short transcripts at the human immunodeficiency virus type 1 long terminal repeat. J Virol. 1997 Feb;71(2):910–917. doi: 10.1128/jvi.71.2.910-917.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Pendergrast P. S., Morrison D., Tansey W. P., Hernandez N. Mutations in the carboxy-terminal domain of TBP affect the synthesis of human immunodeficiency virus type 1 full-length and short transcripts similarly. J Virol. 1996 Aug;70(8):5025–5034. doi: 10.1128/jvi.70.8.5025-5034.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Plet A., Eick D., Blanchard J. M. Elongation and premature termination of transcripts initiated from c-fos and c-myc promoters show dissimilar patterns. Oncogene. 1995 Jan 19;10(2):319–328. [PubMed] [Google Scholar]
  54. Rasmussen E. B., Lis J. T. Short transcripts of the ternary complex provide insight into RNA polymerase II elongational pausing. J Mol Biol. 1995 Oct 6;252(5):522–535. doi: 10.1006/jmbi.1995.0517. [DOI] [PubMed] [Google Scholar]
  55. Ratnasabapathy R., Sheldon M., Johal L., Hernandez N. The HIV-1 long terminal repeat contains an unusual element that induces the synthesis of short RNAs from various mRNA and snRNA promoters. Genes Dev. 1990 Dec;4(12A):2061–2074. doi: 10.1101/gad.4.12a.2061. [DOI] [PubMed] [Google Scholar]
  56. Rickles R. J., Darrow A. L., Strickland S. Differentiation-responsive elements in the 5' region of the mouse tissue plasminogen activator gene confer two-stage regulation by retinoic acid and cyclic AMP in teratocarcinoma cells. Mol Cell Biol. 1989 Apr;9(4):1691–1704. doi: 10.1128/mcb.9.4.1691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Roberts S., Purton T., Bentley D. L. A protein-binding site in the c-myc promoter functions as a terminator of RNA polymerase II transcription. Genes Dev. 1992 Aug;6(8):1562–1574. doi: 10.1101/gad.6.8.1562. [DOI] [PubMed] [Google Scholar]
  58. Rougvie A. E., Lis J. T. Postinitiation transcriptional control in Drosophila melanogaster. Mol Cell Biol. 1990 Nov;10(11):6041–6045. doi: 10.1128/mcb.10.11.6041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Rougvie A. E., Lis J. T. The RNA polymerase II molecule at the 5' end of the uninduced hsp70 gene of D. melanogaster is transcriptionally engaged. Cell. 1988 Sep 9;54(6):795–804. doi: 10.1016/s0092-8674(88)91087-2. [DOI] [PubMed] [Google Scholar]
  60. Roy A. L., Meisterernst M., Pognonec P., Roeder R. G. Cooperative interaction of an initiator-binding transcription initiation factor and the helix-loop-helix activator USF. Nature. 1991 Nov 21;354(6350):245–248. doi: 10.1038/354245a0. [DOI] [PubMed] [Google Scholar]
  61. Ruppert S. M., McCulloch V., Meyer M., Bautista C., Falkowski M., Stunnenberg H. G., Hernandez N. Monoclonal antibodies directed against the amino-terminal domain of human TBP cross-react with TBP from other species. Hybridoma. 1996 Feb;15(1):55–68. doi: 10.1089/hyb.1996.15.55. [DOI] [PubMed] [Google Scholar]
  62. Selby M. J., Bain E. S., Luciw P. A., Peterlin B. M. Structure, sequence, and position of the stem-loop in tar determine transcriptional elongation by tat through the HIV-1 long terminal repeat. Genes Dev. 1989 Apr;3(4):547–558. doi: 10.1101/gad.3.4.547. [DOI] [PubMed] [Google Scholar]
  63. Sheldon M., Ratnasabapathy R., Hernandez N. Characterization of the inducer of short transcripts, a human immunodeficiency virus type 1 transcriptional element that activates the synthesis of short RNAs. Mol Cell Biol. 1993 Feb;13(2):1251–1263. doi: 10.1128/mcb.13.2.1251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  64. Sheridan P. L., Schorpp M., Voz M. L., Jones K. A. Cloning of an SNF2/SWI2-related protein that binds specifically to the SPH motifs of the SV40 enhancer and to the HIV-1 promoter. J Biol Chem. 1995 Mar 3;270(9):4575–4587. doi: 10.1074/jbc.270.9.4575. [DOI] [PubMed] [Google Scholar]
  65. Smale S. T., Baltimore D. The "initiator" as a transcription control element. Cell. 1989 Apr 7;57(1):103–113. doi: 10.1016/0092-8674(89)90176-1. [DOI] [PubMed] [Google Scholar]
  66. Soeller W. C., Oh C. E., Kornberg T. B. Isolation of cDNAs encoding the Drosophila GAGA transcription factor. Mol Cell Biol. 1993 Dec;13(12):7961–7970. doi: 10.1128/mcb.13.12.7961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  67. Spencer C. A., Groudine M. Transcription elongation and eukaryotic gene regulation. Oncogene. 1990 Jun;5(6):777–785. [PubMed] [Google Scholar]
  68. Strobl L. J., Eick D. Hold back of RNA polymerase II at the transcription start site mediates down-regulation of c-myc in vivo. EMBO J. 1992 Sep;11(9):3307–3314. doi: 10.1002/j.1460-2075.1992.tb05409.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  69. Sáfrány G., Perry R. P. The relative contributions of various transcription factors to the overall promoter strength of the mouse ribosomal protein L30 gene. Eur J Biochem. 1995 Jun 15;230(3):1066–1072. doi: 10.1111/j.1432-1033.1995.tb20657.x. [DOI] [PubMed] [Google Scholar]
  70. Toohey M. G., Jones K. A. In vitro formation of short RNA polymerase II transcripts that terminate within the HIV-1 and HIV-2 promoter-proximal downstream regions. Genes Dev. 1989 Mar;3(3):265–282. doi: 10.1101/gad.3.3.265. [DOI] [PubMed] [Google Scholar]
  71. Yankulov K., Blau J., Purton T., Roberts S., Bentley D. L. Transcriptional elongation by RNA polymerase II is stimulated by transactivators. Cell. 1994 Jun 3;77(5):749–759. doi: 10.1016/0092-8674(94)90058-2. [DOI] [PubMed] [Google Scholar]
  72. Yoon J. B., Li G., Roeder R. G. Characterization of a family of related cellular transcription factors which can modulate human immunodeficiency virus type 1 transcription in vitro. Mol Cell Biol. 1994 Mar;14(3):1776–1785. doi: 10.1128/mcb.14.3.1776. [DOI] [PMC free article] [PubMed] [Google Scholar]
  73. Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis of DNA fragments cloned into M13 vectors. Methods Enzymol. 1983;100:468–500. doi: 10.1016/0076-6879(83)00074-9. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES