| | |  | | | | Record from Web of Science® | | | | | | |
| FACS-optimized mutants of the green fluorescent protein (GFP) |
|
|
| Author(s): Cormack BP, Valdivia RH, Falkow S |
| Source: GENE Volume: 173 Issue: 1 Pages: 33-38 Published: JUL 1 1996 |
| Times Cited: 1,508 References: 10 |
| Abstract: We have constructed a library in Escherichia coli of mutant gfp genes (encoding green fluorescent protein, GFP) expressed from a tightly regulated inducible promoter. We introduced random amino acid (aa) substitutions in the twenty aa flanking the chromophore Ser-Tyr-Gly sequence at aa 65-67. We then used fluorescence-activated cell sorting (FAGS) to select variants of GFP that fluoresce between 20- and 35-fold more intensely than wild type (wt), when excited at 488 nm. Sequence analysis reveals three classes of aa substitutions in GFP. All three classes of mutant proteins have highly shifted excitation maxima, In addition, when produced in E. coli, the folding of the mutant proteins is more efficient than folding of wt GFP. These two properties contribute to a greatly increased (100-fold) fluorescence intensity, making the mutants useful for a number of applications. |
| Document Type: Proceedings Paper |
| Language: English |
| Reprint Address: Cormack, BP (reprint author), STANFORD UNIV, SCH MED, DEPT MICROBIOL & IMMUNOL, FAIRCHILD D035, STANFORD, CA 94305 USA |
Addresses:
1. NIAID, ROCKY MT LABS, HAMILTON, MT 59840 USA |
| Publisher: ELSEVIER SCIENCE BV, PO BOX 211, 1000 AE AMSTERDAM, NETHERLANDS |
| Subject Category: Genetics & Heredity |
| IDS Number: UY073 |
| ISSN: 0378-1119 |
|
| | | | | | | | | Record from Web of Science® | | | | | | | | | |