Chlorella is an attractive organism for complex recombinant protein production because of its eukaryotic characteristics and low cost for large-scale culture. Protoplasts of C. ellipsoidea were transformed with a vector containing the flounder growth hormone gene (fGH) under the control of the cauliflower mosaic virus 35S promoter, and the phleomycin resistance Sh ble gene under the control of the Chlamydomonas RBCS2 gene promoter. The presence of introduced DNA was first determined by PCR amplification of both the fGH and Sh ble genes from genomic DNA isolated from transformants and fGH protein expression was detected by immunoblot analysis. Over 400 microg of fGH protein expression per one liter culture containing 1 x 10(8) cells/ml was estimated by ELISA. Stable integration of introduced DNA was confirmed by Southern blot analysis of genomic DNA digested with restriction enzymes. The introduced DNA and fGH expression were detected after seven successive transfers in media devoid of phleomycin, but stably remained in the presence of the antibiotic. Flounder fry fed on the transformed Chlorella revealed a 25% growth increase after 30 days of feeding.

An aerobic, non-motile, Gram-negative, orange-pigmented, rod-shaped, astaxanthin-producing marine bacterium was isolated from the Haeundae Coast, Korea. This strain, BC74171 T , produced carotenoids, mainly astaxanthin. All the type strains of the genus Paracoccus were compared with strain BC74171 T using 16S rRNA gene sequence analysis, fatty acid patterns and physiological reaction profiles. Based on the results of these analyses, it is proposed that strain BC74171 T represents a novel species, Paracoccus haeundaensis sp. nov. The type strain is BC74171 T (=KCCM 10460 T =LMG P-21903 T ).Astaxanthin (3,39-dihydroxy-b,b-carotene-4,49-dione) is a carotenoid that is widely distributed in nature and is present in marine animal tissues, including those of red sea bream, salmon and lobster (Fujita et al., 1983;Johnson & An, 1991;Nelis & De Leenheer, 1991). Organisms that produce astaxanthin include the basidiomycetous yeast Phaffia rhodozyma (Miller et al., 1976), the green alga Haematococcus pluvialis (Bubrick, 1991), and the Gramnegative bacteria 'Agrobacterium aurantiacum' (Yokoyama et al., 1994), Paracoccus marcusii (Harker et al., 1998), Paracoccus carotinifaciens (Tsubokura et al., 1999) and Paracococcus sp. strain MBIC 01143 (Misawa et al., 1995).The genus Paracoccus consists of Gram-negative, oxidaseand catalase-positive bacteria that show substantial metabolic versatility. At present, the genus Paracoccus includes 17 recognized species. Phylogenetically, the genus belongs to the a-3 subclass of the Proteobacteria. In this study, results of a taxonomic study and a phylogenetic analysis based on 16S rRNA gene sequence comparisons are described. Data show that a newly isolated marine strain, BC74171 T , should be classified as a representative of a novel species within the genus Paracoccus. The name Paracoccus haeundaensis sp. nov. is proposed; the type strain is BC74171 T (=KCCM 10460 T =LMG P-21903 T ) and its characteristics are described.An orange-pigmented, astaxanthin-producing bacterial strain, BC74171 T , was isolated from sea water collected on the Haeundae Coast, Korea. This strain was isolated on nutrient agar medium (Difco) and maintained on PPES-II medium (0?2 % polypeptone, 0?1 % Bacto-yeast extract, 0?1 % Bacto-soytone, 0?1 % Bacto-tryptone, 10 p.p.m. ferric citrate and 3 % NaCl) by serial inoculation. Strain BC74171 T was Gram-negative and rod-shaped. The cells ranged from 0?3 to 0?7 mm in diameter and 0?8 to 2?5 mm in length. Cells were non-motile and non-spore-forming. Colonies on agar were smooth, flat and bright orange in colour.Cultured cells were suspended in 0?1 M phosphate buffer (pH 7?2). Cells were fixed with 2 % glutaraldehyde, washed with 0?05 M cacodylate buffer and post-fixed with 1 % osmium tetroxide. Fixed cells were dehydrated in ethanol and the ethanol was replaced with propylene oxide before embedding in Epon resin and sectioning with an ultramicrotome. Sections were examined with a JEM 1200EX-II transmission electron microscope ( Fig. 1; see also the transmission electron mic...

Eight bacterial strains identified as P1, P2, Y1, Y2, W1, W2, G, and R were isolated from a longterm laboratory culture of the green alga Chlorella ellipsoidea. Although it is unknown how these bacterial strains have been maintained with the C. ellipsoidea culture, all appeared to promote the growth of C. ellipsoidea. Co-inoculation of each bacterial strain with C. ellipsoidea resulted in 0.5-3 times greater algal growth than that of C. ellipsoidea alone. The most effective bacterium (i.e., strain P1) was selected and further characterized. Biochemical analysis and transmission electron microscopy revealed that strain P1 is closely related to the genus Brevundimonas. Sequence analysis of the 16S rRNA of strain P1 showed 99.9 and 99.4% nucleotide sequence identity to that of B. nasdae and B. vesicularis, respectively. In addition to the growth promotion of C. ellipsoidea by strain P1, the growth of strain P1 was also significantly enhanced by co-culturing with C. ellipsoidea, indicating a symbiotic relationship between the bacterium and alga. Scanning electron microscopy showed the direct adhesion of strain P1 cells to the surface of C. ellipsoidea cells, as well as the development of abundant crinkles on the surface of co-cultured C. ellipsoidea cells.

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