pSURF-2, a modified BAC vector for selective YAC cloning and functional analysis
A modified bacterial artificial chromosome (BAC) vector, pSURF-2, adapted for the selective subcloning of yeast artificial chromosome (YAC) sequences was constructed. DH10B-U, a pyrF derivative of the highly transformable E. coli strain DH10B was also constructed and used for the detection of Ura+ r...
| Publié dans: | BioTechniques. - 1993. - 27(1999), 1 vom: 01. Juli, Seite 164-70, 172, 175 |
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| Auteur principal: | |
| Autres auteurs: | , , , , |
| Format: | Article |
| Langue: | English |
| Publié: |
1999
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| Accès à la collection: | BioTechniques |
| Sujets: | Journal Article Research Support, Non-U.S. Gov't CFTR protein, human Cinnamates DNA, Bacterial DNA, Fungal Genetic Markers Viral Proteins Cystic Fibrosis Transmembrane Conductance Regulator 126880-72-6 plus... |
| Résumé: | A modified bacterial artificial chromosome (BAC) vector, pSURF-2, adapted for the selective subcloning of yeast artificial chromosome (YAC) sequences was constructed. DH10B-U, a pyrF derivative of the highly transformable E. coli strain DH10B was also constructed and used for the detection of Ura+ recombinants carrying DNA linked to YAC right arms. The vector's properties were illustrated in two main ways. (i) An intact 25-kb YAC containing a mouse tyrosinase minigene was cloned into pSURF-2. Appropriately spliced tyrosinase RNA was detected by reverse transcription (RT)-PCR in extracts of cells transiently lipofected with the cloned YAC. (ii) Cells expressing human cystic fibrosis transmembrane conductance regulator (CFTR) from an integrated pSURF-2 recombinant containing a cDNA expression cassette were selected using the hygromycin-resistance (HyTK) marker of the vector and characterized by RT-PCR and immunoprecipitation. The unique I-SceI site and HyTK marker of pSURF-2 are designed to facilitate subsequent functional studies of cloned DNA |
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| Description: | Date Completed 01.09.1999 Date Revised 28.09.2018 published: Print Citation Status MEDLINE |
| ISSN: | 0736-6205 |