OHSU

Instrumentation

Long-read DNA Sequence and Fragment Analysis

Sequence determination and analysis

AB (Life) 3130XL Fluorescence DNA Analyzer

We use an Applied Biosystems 16-capillary 3130xl automated sequence analysis system purchased through our NIH-NCRR shared instrumentation grant (S10-RR020977). It has a fast laser and uses the "POP-7®" multi-use, high-resolution, polymer for fast fluorescent data collection. The system provides automated polymer loading, sample loading, detection & analysis of both DNA sequencing and fragment analysis samples. Ancillary equipment for cycle sequencing is formatted for 96-well plates. We can process 3 x 96-well plates per day. At 800 nts per read that gives a maximum throughput of about 106 bases a week.

We are using the Applied Biosystems "Sequence Analysis Software" v.5.2 with the "KB" basecaller v.1.2.

Salts and EDTA can mess up the Polymerase activity. Choose a template purification protocol specifically for fluorescent sequencing, attention to detail is critical.

We can perform data assembly and analysis for large sequencing and fragment analysis (SNP, AFLP, etc). Call or email regarding help with bioinformatics and research database development.


Custom Oligo and siRNA Synthesis

Solid-phase synthesis chemistry cycle

ABI 394 Automated Solid-phase Synthesizers

Synthetic oligos are single stranded with hydroxyl groups (not phosphates) at both termini. To form a duplex with the complement: simply add equimolar amounts to the your buffered solution, heat to well over the Tm, then allow to slowly cool to room temperature on the bench. Store cold.

We have two 4-column ABI 394 solid-phase DNA synthesizers. They each have 4 additional reagent ports for incorporating multiple labels, 5'-phosphate, biotin, and numerous base analogs. It is also able to synthesize RNA and mixed RNA and DNA. We are a very cost effective source for synthetic siRNA molecules.