Vector construction. The blunt-ended 4.1 kb Bam HI and Sac I fragment of pQB2, 39 was ligated with the kanamycin resistance fragment ( kan ) that had been amplified from pHSG299, 40 by using the oligo DNA primers: 5′-CGGCCCTGAGGCCTTCAACTCAGCAAAAGTTCG-3′ and 5′-GGCCATATAGGCCTGAATGGCGAATGCGATTTATTC-3′ to create pQA2-Km. To prepare a fragment having a lacI ^q and terminator, pGEX-2TK, 41 was digested by Aat II and Eam I and blunt ended. The resulting lacI ^q fragment was ligated with blunt-ended Bam HI chloramphenicol resistance fragment of pNK2884, 42 to create pCX2TK1. Blunt-ended Aat II and Nhe I fragment of pQA2-Km and Sma I– Fsp I blunt-end fragment was ligated to create pCA21. Original GFP is heat labile and have weak or no fluorescence at 37°C or higher temperature. To replace the original GFP by the heat tolerant mutant form, the Stu I– Sma I fragment of pCA21 was ligated with PCR-amplified GFP fragment from pGFPgcn4, 28 by oligo DNA primers having Not I restriction site: 5′-CCTATGCGGCCGCAGTAAAGGAGAAGAACTTTTC-3′ and 5′-TTAGCGGCCGCTTATTTGTATAGTTCATCCATGCC-3′. Not I sites of both primers were designed for removal of GFP after cloning. The replication origin and Histidine tag of the resultant vector, named pCA24N, were derived from pGEX-2TK and pQE31 (parental plasmid of pQB2) respectively.