Chromosome changes in cell lines of the box turtle (Terrapene carolina) grown at two different temperatures.

Abstract

Introduction Cell lines dcrived from poiliilothermic vertebrates such as rainbow trout (Salsl7zo gairdneri), bullfrog (Rmln catesbeiann), fathead minnow (Pirnephnles pronzelns), a marine turtle (Cheloslin nzydns), and box turtle (Terrape17e carolinn) have been established in continuous culture and used for viral and biochemical studies (Wolf and Quimbv, 1962, 1964; Gravell and Malsberger, 1965; kvaddell and Sigel, 1965; Clarlc and I<arzon, 1967a). In using these cell lines as tools for biological research, a lmowledge of their cytogenetic stability is essential. In contrast to long term cell cult~lres of higher vertebrates, there is little information on the chromosome patterns in cell lines of poiltilothermic vertebrate origin. Wolf and Quimby (1962, 1964) found a subdiploid modal number in a cell line of rainbow trout and triploid in a bullfrog line. In cultured cclls of fathead minnow, Gravell and hlalsberger (1965) observed the model chron~oson~e number to be subdiploid at the 20th and 40th passage, and diploid plus one at the 50th passage. Recently a reinvestigation of chromoson~e patterns of cell lines from the bullfrog and tlye fathead minnow bv Levan et 01. (1966) also shoxved modal numbers of triploid and diploid plus one respectively. Waddell and Sigcl (1965) indicated that the cclls of a continuous line from a lnarine turtle retained their normal diploid number up to the 53rd passage. In all these studies, a precise description of the normal diploid patterns or a karyotype analysis of thc cell lines is lacking. The present paper reports numerical and structural chromosome changes observed in nine cell lines and sublines derived from normal tissues of three box turtles. The differential effect on the cliromosomes of cell cultures incubated at ambient ( 2 3 C) and elevated temperature (30°C) is described. T o our Itnowledge, the somatic chromoson~es of this species have not previously been reported. In ordcr to compare the chromosome patterns in cell lines with the normal diploid sct, the liaryotvpe of primary culturcs from two animals is also described. Materials and Methods Cell Czl1tz11.e~: In Junc, 1965, two adult female Terrnpene carolif7n, designated T 4 and TS, werc ltilled and exsanguinated. Hearts, spleens and lungs were ~vashcd separately w-ith amphibian Ringer's solution, minced finely and explanted in plastic flasks (Falcon) and on coverslips in Leighton tubes in Eagle's basal medium plus lo0/; fetal calf serum. Cultures were incubated at room temperature (23°C) or at 30°C. Leighton tube cultures incubated at 30°C were harvested for determination of the normal chromosome constitution at about 10 to 15 davs.