Applications Genomics 2017-02-15T17:18:58+00:00

The DNA molecule

Genomics is the study of an organism’s genome, or genes and their function. Genomics aims to understand the structure of the genome, including the mapping genes and sequencing the DNA. Genomics examines the molecular mechanisms and the interplay of genetic and environmental factors in disease.

Genomics includes:

  • Functional genomics — the characterization of genes and their mRNA and protein products.
  • Structural genomics — the dissection of the architectural features of genes and chromosomes.
  • Comparative genomics — the evolutionary relationships between the genes and proteins of different species.
  • Epigenomics (epigenetics) — DNA methylation patterns, imprinting and DNA packaging.
  • Pharmacogenomics — new biological targets and new ways to design drugs and vaccines.

Dynamic Devices fully automates a large variety of genomic applications that integrate many different products and technologies. By combining different kits, sealers, thermocyclers, plate readers, shakers and incubators virtually all the manual processes can be automated. The listed applications below is just a short list of the possibilities:

  • Large Volume Genomic DNA Purification
  • Total RNA Isolation from cells
  • Nucleic Acid Purification
  • DNA / RNA Quantification
  • Individual DNA Normalization
  • Sub Microliter PCR Setup
  • 96 and 384 Plate Format PCR Cleanup
  • cDNA Synthesis

D2’s Volume Verified Pipetting (VVP) offers sample transfer volumetric validation within applications that deal with complex sample matrixes. Tissue, cells, serum, plant extracts and even forensics field sample collection utensils, like swabs, create pipetting issues for liquid handlers without the ability to monitor, control and recover from clogging. VVP not only offers the recovery of complex pipetting matrixes but actively accommodates for sample viscosity and temperature automatically eliminating the need to develop complex liquid classes for every type of sample matrix.

Independent (i8) and fixed-8 (f8) VVP tip tools and, the flagship, high throughput 96 Channel VVP Pipetting Tool offer the same type of air based verified volume transfers with full liquid handling diagnostics offering independent monitoring AND independent recovery from EACH tip in the head.

Nucleic Acid Purification – Large Volume & Plasma / Liquid Biopsy Samples

Large Volume Purification System Replacing KingFisher Process

Dynamic Devices Fully Automated Plasma & Liquid Biopsy Processing

Large Volume DNA Mag Bead Extraction
As automation simplifies DNA purification from larger volumes tax the volume ranges of most mainstream automation systems. Dynamic Devices introduces its 24 channel extraction tool with 5mL pipetting tips in conjunction with Click-Bio’s large volume extraction block technology.

24VVP – 24 Independent Volumes
The 24 VVP Pipetting Tool now allows high throughput DNA purification by having the ability to aspirate or dispense 24 different volumes using 5mL tips. With a Lynx LM1200, the 24 VVP Pipetting Tool and head microplate gripper tool hundreds of samples per run may be processed in a fully automated method.

Process 30-50mL Liquid Biopsy Samples
Combining this configuration with the Clickbio’s 12 well large volume extraction plates, 2 tips may process a sample simultaneously giving a 10mL working volume.

Summary of Features:
• Runs 12 or 24 samples simultaneously
• ClickBio block can be moved by gripper to shaker and mag plate as entire unit, no individual tubes
• Processing block hold 30-50mL of liquid
• Reads bar codes and reports every volume transfer
• Liquid level detection and clog/clog detection with VVP technology
• Delivers final elution in 96 SBS standard microplate

SPECIAL HARDWARE NOTE: Clickbio X-Block 

Nucleic Acid Purification

Fully Automated High-throughput Nucleic Acid Purification System Using Magnetic Bead Technology

Lynx DNA Mag BEad Purification System

Dynamic Devices Fully Automated Mag Bead System

Using the Lynx liquid handling robot system comprising of an Dynamic Devices in-deck 6 position Magnetic Bead Purification System prepares 6 plates at single time with no plate movement or gripper tool required. Place the purification plates into the deck position and magnets are brought up from the bottom to pull the beads to the sides of the plate during washing routines. During capture, washing and elution the 96 channel head uses up to 1250uL tips to mix the samples, add or remove liquid from each well and then add elution buffer to remove DNA from the beads and transfer to a clean destination 96 or 384 well microplate.

The Lynx liquid handling robot precisely dispenses low volumes of DNA template and then combines the primer for the PCR protocols. This low volume dispensing permits the scale reduction of the reagents while maintaining kinetics and performance.

SPECIAL HARDWARE NOTE: In Deck 6 Position Magnet Separation System

Fully Automated High-throughput Nucleic Acid Purification System Using Solid Phase Extraction (SPE) Technology

invirtogen plate1

spe system front

Dynamic Devices Fully Automated SPE Vacuum System

Using this liquid handling system comprising of an Lynx Liquid Handler with a 96 head, a gripper tool and a Dynamic Devices 6 position vacuum manifold kit. The compact and flexible Lynx SPE Vacuum System is a fully integrated system to carry out solid phase extraction (SPE). The fully automated liquid handling robot works with 96 well format SPE separation plates that may take the place of the standard manual syringe type SPE formats.

  1. Prepare the SPE matrix with a reagent
  2. Add sample with contamination free disposable tips
  3. Vacuum sample through matrix where excess liquid is drained to a waste container
  4. Move matrix plate over a clean vacuum position with a clean collection plate
  5. Add elution buffer
  6. Vacuum purified sample into collection plate

SPECIAL APP NOTE: Positive Pressure SPE Purification

The 96 VVP & Fixed-8 VVP Pipetting Tools may also be configured to apply positive pressure and monitor the flow rate of the capture and washing protocols. This special application assure there is no ‘blow by’ from the first samples drying out and a large percentage of the vacuum being pulled down the already dry wells.

Purifying Complex Samples – DNA from Whole Blood & Plant/Seed Homogenate 

96-vvp-clot-detection-correction-sm

D2’s Volume Verified Pipetting (VVP) offers sample transfer volumetric validation within applications that deal with complex sample matrixes. Tissue, cells, serum, plant extracts and even forensics field sample collection utensils, like swabs, create pipetting issues for liquid handlers without the ability to monitor, control and recover from clogging. VVP not only offers the recovery of complex pipetting matrixes but actively accommodates for sample viscosity and temperature automatically eliminating the need to develop complex liquid classes for every type of sample matrix.

VVP – Colt/Clog detection and on-the-fly correction
One of the most sought after functionalities in liquid handling robotics is the ability to run individual channels within a high throughput head like the 96 VVP Pipetting Tool. Now, when pipetting errors are encountered by a channel, the aspiration is immediately stopped for just the erred channel while the reset of the samples are aspirated correctly. The head may then clear the erred channels above the liquid and dip the head again into the samples to re-try the aspiration to correct for the error on-the-fly.

DNA Normalization

By calculating the concentration of DNA with a plate reader and knowing the volume of the DNA sample, the total amount of DNA purified can be quantified. In some automated processes using a liquid handling robot, the amount of DNA in each well of a 96 or 384 well microplate is standardized / normalized, to optimize a test or gel to run more accurately. To normalize DNA, the DNA sample absorbance from the reader are examined from a microplate and a spreadsheet or database can calculate the amount of water to dilute each individual sample with in order to get all the samples to the same concentration.

The 96 VVP Pipetting Tool now allows high throughput DNA normalization by having the ability to dispense 96 different volumes in a single dispense. With a Lynx LM1200 and the 96 VVP Pipetting Tool hundreds of plates per day may be normalized with a standard run of 10 plates taking 5 minutes to normalize using a single set of 1250uL tips.

VVP Residual Volume Data

DNA normalization is now optimized using a 96 VVP Head. With the ability of independently controlling each tip with individual volumes and volume verification, an entire 96 well DNA sample plate may be diluted and mixed in a single step. By using a worklist protocol method, it is now possible to take a plate containing various concentrations of DNA. An electronic file containing each wells DNA concentration calculated from a plate reader at 260/280nm, volume and concentration are used to calculate a corresponding amount of ddi water to dilute all the DAN samples to a standard concentration.

The Lynx liquid handling robot’s 96 VVP Pipetting Tool can be used to dilute DNA plates for normalization in multiple high throughput ways:

  1. An entire 96 well compound plate can be diluted with different volumes in one aspirate and dispense step.
  2. A 384 well compound plate may be diluted in 5 steps with a single aspiration and 4 dispenses since the head may multi-pipet different volumes over multiple plates.

In addition, the Fixed-8 (f8) VVP Pipetting Tool tips may also perform any of the above tasks with the same performance and validated performance in multi-dispensing mode.

PCR Preparation

384SV w 3ul under sdie view_KBK8973

The loading and transferring low volumes or 0.5 to 1 µL of DNA and Nucleic Acid samples for amplification is an easy and straightforward task for the Lynx Series Liquid Handling Robot. Once a sample of DNA is isolated, through processes like solid phase or magnetic beads, amplification may take place in a variety of different formats. The transfer of 96 or 384 samples into a thermocycler plate takes place right on the deck and can even be transferred into a plate sealer for the addition of an aluminum foil seal to prevent evaporation in the heating / cooling process.

Additionally, the PCR preparation can automatically follow a liquid handling robot set up for DNA purification and, if the cycle time are short, plates can be loaded directly into a thermocycler or RT-PCR machine.

Automated Expression Cloning

In expression cloning, expression vectors of a particular interest are used to generate a library of clones that each express only one protein. These expression vectors, or DNA segments attached to plasmids, are transfected into cells that need to be grown and maintained.

  • Automation Liquid Handling Robot Platform & Worktable
  • Cylindrical Robotic Manipulator & Servo Gripper
  • Sixty Position Hotel Micro Plate & Lid Storage
  • Single Position Delidder Station
  • Cytomat 2C Cell Based Incubator
  • Two – MicroFlo Reagent Dispensers
  • Single Position Micro Plate Orbital Shaker
  • HEPA Filtered Containment Hood

Fully Automated Expression Cloning System Capacity & Protocol

The liquid handling robot system configuration is designed to automate the complete walk away process of up to 25 384-Well cell based assay plates and corresponding required disposables. The complete automated process for the 384-Well DNA plates includes the addition of the FuGene/OptiMEM reagent, shaking mix step and 40 – 45 RT incubation followed by the addition of the cell suspension. The shelf life of the mixed FuGene/OptiMEM reagent is established at only 45 minutes, but this is no longer an issue as the throughput for the entire process will be high. The limitation to the total throughput for the process is limited by the 40 – 45 RT incubation step. All 25 384-well DNA plates will be processed and into the 40 – 45 RT incubation step before the first plate has completed this incubation step. Once this incubation time has completed for plate 1, 20 µl of cell suspension is dispensed. The process time for the cell suspension step will be almost identical to the FuGene/OptiMEM step, thus providing a very consistent assay time between one plate to the next. The overall process time is estimated at 71 minutes for the complete process for all 25 plates.

Fully Automated Expression Cloning Liquid Handling Robot System Worktable Layout

expression cloning plate prep system 1 lg px600
Each 96 well DNA plate will have a new tip box associated with it and used for both transfer steps of the source DNA. This includes the transfer of each DNA plate to the 96 well reaction plate as well as the subsequent transfer from the reaction plate to the 384 well cell based assay plate. Likewise, the FuGene/OptiMEM reagent will have a dedicated set of disposable tips that are dedicated to each transfer of the reagent. During the transfer of this reagent, no contact will be made with the destination plate during the dispense steps.

Automated Post PCR Cleanup

Since most added unincorporated primers and dNTP’s may not be used up during the amplification process, a cleanup may be necessary before the samples are carried on to the next step.  This cleanup may take place in different forms such as spin column with a silica membrane or, the best solution for automation, a filter based plate 96 or 384 format.  These plates can use a size-exclusion membrane and vacuum filtration for a single step process that fits great into an Oasis liquid handling robot equipped with a 96 well head and gripper tool.

For improved productivity, 96 samples can be processed in less than 15 minutes with most standard kits available on the market.  The system transfers the samples onto the filtration plates, pulls through the leftover materials to waste and then places a clean collection plate under the filter plate and adds elution buffer.  The amplification product is then pulled the membrane with the vacuum box and the concentrated and purified DNA fragments are collected for such applications as sequencing, ligation and microarrays.
In addition, if cleanup is required from an agarose gel, the same configuration Oasis liquid handling robot can process these same samples after the gel is digested.

DNA Quantification

A simple microplate OD or fluorescent plate reader is used to quantify the concentration of DNA, combined with the volume associated with the sample, a quantity of DNA can be calculated.  By integrating an automation friendly plate reader (as described in the integration section) with a liquid handling robot, quantification of DNA may be calculated ands stored on-line to an automated system.  The sample concentrations may then be standardized, or normalized, with a liquid handling robot by adding a calculated amount of water to each sample.  This process is necessary when a consistent concentration of DNA is required for PCR setup or running a gel.