Isolation and culture of protoplasts from high anthocyanin-producing callus of sweet potato

Optimum conditions for the isolation and culture of protoplasts from high anthocyanin-producing callus of sweet potato (Ipomoea batatas) were investigated. Growth phase of callus and the ratio of callus-enzyme solution affected the yield of protoplasts. Composition of the medium and protoplast density were examined for protoplast culture.Small colonies were regenerated from the protoplasts. Upon transfer to light a high amount of anthocyanin was accumulated in these colonies.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - MES 2-(N-morpholino)-ethanesulfonic acid     

Influence of adenosine 3':5'-cyclic monophosphate analogues on testicular organization of fetal mouse gonads in vitro

Gonadal primordia, isolated from fetal mice on the 11th or 12th day of gestation, differentiated in vitro into morphologically distinct testes or ovaries after 7 days in culture. The addition of cAMP analogues into culture media prevented the differentiation of testis cords. Histological examination indicated that the basement membranes of testis cords disintegrated after treatment with cAMP analogues, while development of germ cells and Leydig cells appeared to be unaffected. Fetal testes in culture secreted testosterone which increased following addition of dibutyryl-cAMP (Bt2 c-AMP). Primordial germ cells reached prespermatogonial stage in the presence or absence of Bt2 cAMP, suggesting that progressive differentiation of primordial germ cells is independent of testis cord organization. The Bt2 cAMP-treated explants resumed testicular development after transplantation into a site beneath the kidney capsules of adult mice, although the inhibitory effect appeared irreversible in vitro. The testicular organization-preventing effect of cAMP analogues was mimicked by prostaglandins or forskolin, which are known to stimulate adenylate cyclase. The inhibitory effect of either cAMP analogues or prostaglandins was potentiated when added in combination with phosphodiesterase inhibitors. The present results suggest that increase of intracellular cAMP prevents the development of basement membrane and the assembly of cells to form testicular structures.     

Deletions in a recombinant retrovirus genome associated with its expression in embryonal carcinoma cells.

We analyzed embryonal carcinoma cell lines infected with a recombinant Moloney murine leukemia virus. Lines that carried but did not express the neo gene retained a provirus of LTR-gag-pol-neo-LTR, where LTR is a long terminal repeat, whereas all G418-resistant lines deleted regions that included the primer binding site and the splicing donor site. This suggested the presence of multiple inhibitory elements.     

Genetic variation in the Y chromosome and sex-biased DNA methylation in somatic cells in the mouse

Mammalian Genome - Several lines of evidence suggest that the presence of the Y chromosome influences DNA methylation of autosomal loci. To better understand the impact of the Y chromosome on...     

Synthesis of bacteriophage phiC DNA in dna mutants of Esherichia coli.

Host dna functions involved in the replication of microvirid phage phiC DNA were investigated in vivo. Although growth of this phage was markedly inhibited even at 35-37 degrees C even in dna+ host, conversion of the infecting single-stranded DNA into the double-stranded parental replicative form (stage I synthesis) occurred normally at 43 degrees C in dna+, dnaA, dnaB, dnaC(D), and dnaE cells. In dnaG mutant, the stage I synthesis was severely inhibited at 43 degrees C but not at 30 degrees C. The stage I replication of phiC DNA was clearly thermosensitive in dnaZ cells incubated in nutrient broth. In Tris-casamino acids-glucose medium, however, the dnaZ mutant sufficiently supported synthesis of the parental replicative form. At 43 degrees C, synthesis of the progeny replicative form DNA (stage II replication) was significantly inhibited even in dna+ cells and was nearly completely blocked in dnaB or dnaC(D) mutant. At 37 degrees C, the stage II replication proceeded normally in dna+ bacteria.     

Replication of bacteriophage G13 DNA in dna mutants of Escherichia coli.

Host functions required for replication of microvirid phage G13 DNA were investigated in vivo, using thermosensitive dna mutants of Escherichia coli. In dna+ bacteria, conversion of viral single-stranded DNA into double-stranded replicative form (stage I synthesis) was resistant to 150 microgram/ml of chloramphenicol or 200 microgram/ml of rifampicin. Although multiplication of G13 phage was severely inhibited at 42--43 degrees C even in dna+ host, considerable amount of parental replicative form was synthesized at 43 degrees C in dna+, dnaA or dnaE bacteria. In dnaB and dnaG mutants, however, synthesis of parental replicative form was severely inhibited at the restrictive temperature. Interestingly enough, stage I replication of G13 DNA was, unlike that of phiX174, dependent on host dnaC(D) function. Moreover, the stage I synthesis of G13 DNA in dnaZ was thermosensitive in nutrient broth but not in Tris/casamino acids/glucose medium. In contrast with the stage I replication, synthesis of G13 progeny replicative form was remarkably thermosensitive even in dna+ or dnA cells.