Project description:cDNA library sequencing in the midgut, mixed segment and first proctodeal segment of worker termites in Nasutitermes takasagoensis by 454 GS Junior pyrosequencing
Project description:Contaminated aquifer (Dusseldorf-Flinger, Germany) templates extracted from 5 sediment depths ranging between 6.4 and 8.4 m below ground and over 3 years of sampling were amplified for amplicon pyrosequencing using the primers Ba27f (5’-aga gtt tga tcm tgg ctc ag-3’) and Ba519r (5’- tat tac cgc ggc kgc tg-3’), extended as amplicon fusion primers with respective primer A or B adapters, key sequence and multiplex identifiers (MID) as recommended by 454/Roche. Amplicons were purified and pooled as specified by the manufacturer. Emulsion PCR (emPCR), purification of DNA-enriched beads and sequencing run were performed following protocols and using a 2nd generation pyrosequencer (454 GS FLX Titanium, Roche) as recommended by the developer. Quality filtering of the pyrosequencing reads was performed using the automatic amplicon pipeline of the GS Run Processor (Roche), with a slight modification concerning the valley filter (vfScanAllFlows false instead of TiOnly) to extract the sequences. Demultiplexed raw reads were furhter trimmed for quality and lenght (>250 bp).
Project description:Phylogeny and the distribution of symbiotic bacteria in the mixed segment of the wood-eating termite Nasutitermes takasagoensis (Shiraki) were studied. Bacterial 16S rRNA genes (rDNA) were amplified from the mixed segment of the gut by PCR, and two kinds of sequences were identified. The phylogenetic tree was constructed by neighbor-joining and maximum parsimony methods to identify symbionts harbored in the mixed segment. They are classified as low-G+C-content gram-positive bacteria and are most closely related to the genus Clostridium. The distribution of these bacteria throughout the whole gut was examined by PCR using specific primers, which suggested that they are confined to the mixed segment despite the presence of bacteria throughout the gut. In situ hybridization indicated that the symbiotic bacteria were localized to the ectoperitrophic space between the midgut wall and the peritrophic membrane in the mixed segment. Electron microscopy revealed the close association between these bacteria and the mesenteric epithelium, suggesting that they have some interactions with the gut tissue of termites.
| S-EPMC101473 | biostudies-literature
Project description:Pyrosequencing Analysis of LaborSaving211 using GS FLX 454
| PRJEB3327 | ENA
Project description:Pyrosequencing Analysis of BlackClust using GS FLX 454
