Faculty of Science - Leading creativity and innovation in the sciences


DNA Sequencing Centre


Applied Biosystems 3130XL Genetic Analyzer


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The 3130XL is a DNA sequencing machine that analyses DNA samples via sanger sequencing (determines the sequence of specific DNA) and genotyping (for sizing of specific DNA fragments) using capillary electrophoresis coupled with fluorescent dye technology.

Service

The DNA sequencing centre offers the 3130XL as a service to UoA and external clients with a number of different options available.

 

Sanger sequencing options available:

 

Full sequencing service:

The Facility undertakes both the fluorescent chemical reaction and electrophoresis run.

 

Capillary electrophoresis service:

The client does the fluorescent chemical reaction and the Facility runs the electrophoresis.

 

Genotyping options available:

Genotyping Internal lane standard size ladder options:

Rox (Red dye) ladder 400HD / 500HD


For the dye selection combination 6 fam/hex/ned.

Liz (Orange dye) ladder 600 /1200 


For the dye selection combination 6 fam/vic/ned/pet.

Techniques

The 3130XL has 16 capillaries that process samples in batches of 16 and can process 96 well or 384 well trays. The samples are processed on the 3130XL using 50cm capillaries and POP 7 polymer.

Sanger sequencing can be used for a number of applications:

De Novo sequencing, Mitochondrial sequencing, Fast resequencing, Bac End Fingerprinting.

Next-Gen sequencing confirmation, and heterozygote detection.

All Sanger sequencing reactions at the Facility are carried out using Applied Biosystems BigDye version 3.1 terminator chemistry on Applied Biosystems 9700 Gold Block thermal cyclers. Cleanup of dye terminators uses CleanSeq magnetic bead technology followed by electrophoresis and analysis on the 3130XL.

Genotyping automates DNA fragment analysis using internal-lane standards and can be used to detect genetic variation. This analysis can size PCR products and accurately analyse molecular lengths in the 50-1000 nucleotide size range.  Size ladder and Hi Di formamide are added to the samples, heat shocked and ice quenched followed by fragment separation and analysis on the 3130XL.

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Agilent Bioanalyzer 2100


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The Agilent Bioanalyzer 2100 is used to determine the quality and quantity of RNA/DNA samples. Twelve samples are loaded onto one chip and run on the machine via electrophoresis. Clients run their own samples using their own kits, or those supplied by the Facility.

Application kits available at the Facility are Agilent DNA High Sensitivity kits and Agilent RNA Nano kits.  

 

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Applied Biosystems 7900HT Sequence Detection System


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This machine uses fluorescence-based PCR (polymerase chain reaction) chemistries that allow for quantitative and qualitative detection of nucleic acid sequences in real time using 384 well trays

Service

Clients run the 7900HT themselves allowing flexibility with project design and time.   

Techniques

qPCR has a number of applications including gene expression analysis; miRNA analysis and analysis of genomic variations using SNP & CNV genotyping.

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Nanodrop and Implen Spectrophometers


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Nanodrops and Implen instruments are low volume spectrophotometers (nanophotometers) that do not require a cuvette. These can be used to measure the quantity of RNA/DNA and protein using only 1 µl of your product. The Facility has two nanophotometers that can measure single samples (Implen N60) and one instrument that has eight wells (Nanodrop 8000).

 

Qubit Fluorescent Spectrophotometer


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The Qubit is used to quantify DNA/RNA and proteins and uses fluorescent dyes for accurate readings. The Facility stocks reagents and tubes.

 

Corbett 384 well PCR machines


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The centre has two machines used for PCR (polymerase chain reaction).  The machines allow cycles of different temperatures in automated programmed steps that make multiple copies of a targeted segment of DNA.  They are 384 well tray sample format. 

 

Applied Biosystems 9700 Thermal Cycler Gold Block - 96 well PCR machines


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The centre has six machines used for PCR (polymerase chain reaction).  The machines allow cycles of different temperatures in automated programmed steps that make multiple copies of a targeted segment of DNA. They are 96 well tray sample format. 

 

Liquid Handling Robots: Eppendorf epMotion 5075 and Beckman Biomek 4000


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Liquid handling robots are available to clients free of charge.  The robots enable the client to free up their time, reduces human pipetting errors and reduces repetitive strain injury. 

 

Our people


Please contact Kristine Boxen for more information or assistance.

Kristine Boxen is the Lab Manager in the DNA Sequencing Centre with 20 years experience in the Genomics field.  

See, Selected Publications