Selection of a cDNA clone for chicken high-mobility-group 1 (HMG1) protein through its unusually conserved 3′-untranslated region, and improved expression of recombinant HMG1 in Escherichia coli
| Autor: | Keng-Boon Lee, Dawn J Brooks, Jean O. Thomas |
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| Rok vydání: | 1998 |
| Předmět: |
clone (Java method)
Untranslated region DNA Complementary Recombinant Fusion Proteins Molecular Sequence Data Biology Conserved sequence Sequence Homology Nucleic Acid Complementary DNA Escherichia coli Genetics Animals Amino Acid Sequence Cloning Molecular Codon 3' Untranslated Regions Conserved Sequence Base Sequence Sequence Homology Amino Acid cDNA library Accession number (library science) Three prime untranslated region High Mobility Group Proteins Gene Expression Regulation Bacterial General Medicine Hmg protein Molecular biology Chickens Sequence Alignment |
| Zdroj: | Gene. 225:97-105 |
| ISSN: | 0378-1119 |
| DOI: | 10.1016/s0378-1119(98)00542-3 |
| Popis: | Screening of cDNA libraries for the homologous vertebrate proteins high mobility group (HMG) 1 and 2 using DNA probes based on the coding sequences is likely to result in isolation of both HMG1 and HMG2 clones, as well as pseudogenes, which may be transcribed at low levels. However, the 3'-untranslated regions (UTRs) of HMG1 and 2 are quite distinct, and unusually conserved across species. We have used this property to select the true chicken HMG1 cDNA clone from a chicken lymphocyte cDNA library in lambdagt11, using a probe based on the 3'-UTR of rat HMG1 cDNA. The chicken HMG1 cDNA clone is very similar to all the complete HMG1 cDNA clones isolated so far. We suggest that the sequence designated chicken HMG1 in the GenBank Data Library (Accession number D14314) is, in fact, that of HMG2a [and moreover that the recently reported mouse clone (Accession number AF022465), proposed to encode a new HMG protein, HMG4, is also likely to encode an HMG2a, based on the translated amino-acid sequence and 3'-UTR]. We also report much improved expression of intact recombinant HMG1 in Escherichia coli by the use of chloramphenicol rather than ampicillin selection and conditions that limit cell growth. This should be general for all members of the HMG1 (and 2) family which may be toxic to cells (possibly because of the long acidic tail), and may also prove useful in the production of other such proteins. |
| Databáze: | OpenAIRE |
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