To contribute to our understanding of the genome complexity of sugarcane, we undertook a large-scale expressed sequence tag (EST) program. More than 260,000 cDNA clones were partially sequenced from 26 standard cDNA libraries generated from different sugarcane tissues. After the processing of the sequences, 237,954 high-quality ESTs were identified. These ESTs were assembled into 43,141 putative transcripts. Of the assembled sequences, 35.6% presented no matches with existing sequences in public databases. A global analysis of the whole SUCEST data set indicated that 14,409 assembled sequences (33% of the total) contained at least one cDNA clone with a full-length insert. Annotation of the 43,141 assembled sequences associated almost 50% of the putative identified sugarcane genes with protein metabolism, cellular communication/signal transduction, bioenergetics, and stress responses. Inspection of the translated assembled sequences for conserved protein domains revealed 40,821 amino acid sequences with 1415 Pfam domains. Reassembling the consensus sequences of the 43,141 transcripts revealed a 22% redundancy in the first assembling. This indicated that possibly 33,620 unique genes had been identified and indicated that >90% of the sugarcane expressed genes were tagged

The pH-independent hydrolysis of 4-nitrophenyl chloroformate in the presence of aqueous micelles of sodium dodecyl sulfate, sodium dodecylbenzene sulfonate, alkyltrimethylammonium chlorides, alkyldimethylbenzylammonium chlorides (alkyl group = cetyl and dodecyl) and polyoxyethylene (9) nonylphenyl ether was studied spectrophotometrically. The observed rate constants, k obs , decrease in the following order: cationic micelles b bulk water b non-ionic micelles b anionic micelles. Surfactant-substrate association constants, K s , were determined from the dependence of k obs on surfactant concentration, and were found to be only slightly dependent on the charge of the surfactant and, for similarly charged micelles, on the length of their hydrophobic tail. A 1 H NMR study of the solubilization of a model compound, 4-nitrophenyl chloroacetate, showed that all surfactant segments are affected by the solubilizate and the effect is more pronounced toward the middle of the hydrocarbon chain. The average solubilization site of the acetate ester does not depend on the charge of the micelle or the length of the surfactant hydrophobic tail. Micellar effects on observed rate constants are analyzed in terms of a 'medium' effect and an 'electrostatic' effect. The lower microscopic polarity at the reaction site retards the reaction, whereas electrostatic interactions of the polar transition state with the charged interface result in a rate decrease by anionic micelles and a rate enhancement by cationic micelles.

We have sequenced a 2.5-kb DNA fragment of the B-2 DNA puff from the sciarid Rhynchosciara americana and have defined its transcription unit. This puff is active during the formation of the communal cocoon, which is important for successful metamorphosis of this species and coincides with the final cycle of polytenization in its salivary glands. The B-2 polypeptide, together with the products of two other previously characterized DNA puffs, seems to be engaged in an interaction that results in a gradual modification and hardening of the cocoon structure. The B-2 messenger is temporally regulated in apparent coordination with the other puff products. The predicted polypeptide has characteristics similar to polypeptides from previously sequenced DNA puff genes, in particular those from the R. americana C-8 gene and the Bradysia hygida C-4 gene. The cloned sequence of the B-2 puff is differentially amplified in the three gland regions examined, achieving its highest amplification level of approximately fourfold (two extra cycles) in the anterior segment of the gland. The C-3 DNA puff sequence was also found to be differentially amplified in the different gland regions. Implications of the widespread presence of DNA amplification as a form of gene regulation in the Sciaridae are discussed.

The pH-independent hydrolyses of 4-nitrophenyl chloroformate, NPCF and 4-nitrophenyl heptafluorobutyrate, NPFB in aqueous acetonitrile were studied spectrophotometrically from 15 to 45 8C. The binary solvent composition covers water concentrations from 2.349 to 53.207 and from 2.745 to 53.333 mol L À1 for NPCF and NPFB, respectively. For both esters, the dependence of log (k obs ), the observed rate constant, on log [water] is sigmoidal. The approximate kinetic orders with respect to water were found to be 2 and 3 for NPCF and NPFB, respectively. DG 6 ¼ gradually decreases as a function of increasing [water], due to a complex, quasi-mirror image compensation of DH 6 ¼ and DS 6 ¼ ; both parameters increase. The structures of the transition states were probed by a proton inventory study, carried out in the presence of L 2 O mole fractions (L ¼ H or D) of 0.190, 0.540, 0.890 and 0.180, 0.529, 0.890, for NPCF and NPFB, respectively. Plots of observed rate constants versus the atom fraction of deuterium in the solvent curve downward. Cyclic transition state models were fitted to the kinetic data; these models contain the ester and two water molecules (NPCF) or three water molecules (NPFB). Thus, the sigmoidal dependences of log (k obs ) on log [water] are not due to a change in the number of water molecules in the transition states as a function of increasing [water]. The binary solvent mixture is micro-heterogeneous; there exists two ''micro-domains,'' one consists predominantly of coordinated water molecules, the other consists mostly of acetonitrile hydrogen-bonded to water molecules. NPCF is 232 times more soluble in water than NPFB. That is, the former ester is dissolved in the outer, more polar periphery of these micro-domains whereas the more hydrophobic NPFB is dissolved in their inner, less polar interiors. This conclusion is corroborated by comparing the dependence on log [water] of log [k obs ], and of E T , the empirical solvent polarity parameter, as measured by solvatochromic probes of increasing hydrophobicity. a The rate constants reported are the mean of at least triplicate runs. The relative standard deviation between k obs of a triplicate, that is, ((standard deviation/ k obs )100) was 1.0%.