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Services – Self-Service Instruments


• Client PC Workstation (networked)

• SmartCheck (Pipette QC & Training)

• qPCR – ABI QuantStudio·6

• PCR (standard) – Veriti thermocycler

• Qubit 4.0 (DNA/RNA/Protein Quantitation)

• Diagenode Bioruptor NGS (DNA shearing)

• ThermoFisher SPD1010 SpeedVac

• Typhoon 8600 Variable Mode Imager

• Eppendorf Centrifuge 5810R

• Victor·3 Multilabel Counter


Suspension of Self-Service

  • Due to a lack of Service Agreements to cover instrument repairs, only Core staff may operate the instruments in the Core.
  • Thus, all samples for Biosci-Booked (Genomics) requests will be transferred to the Core for processing; for complete details, please see Operations during Self-Service Suspension.
  • This policy will be revisited when, and if, Service Agreements are re-instated on Core instrumentation.
  • Instrument Access

    Nearly all of these instruments are accessed only by advanced reservations through the online Biosci-Booked (Genomics) system; however, no reimbursement charges are assessed for their use. Priority users include LSU faculty in the College of Science, as well as their staff and graduate students. Requests by other members of the University (for access to the self-service instruments) will be considered on a case-by-case basis.

    Client PC Workstation

    Client computer station Advance Reservations, made through the Biosci-Booked (Genomics), are required for use of this computational resource (Computer-Workstation.jpg) in the Genomics Core. The computer has a variety of genomics-related software (see List) with associated User Guides, as well as standard Microsoft Office software. For descriptions of some of the ABI software, please see Data Analysis on the 'Services' page for the ABI 3130xl platform. Suggestions for additional 'genomics' software are welcome.

    The Client PC Workstation is on LSU's network, and remote access to this resource requires the computer name (lsa-genomicslab), inclusion on the computer's list of approved remote access clients, and registration in Biosci-Booked (Genomics). Clients must adhere strictly to the schedule in Biosci-Booked (Genomics); otherwise, analyses might be lost when they log in while someone else is already using the computer. When accessing this resource from off-campus, your computer must have the LSU-approved VPN (GlobalProtect VPN, as of late 2020) installed and connected. Finally, clients must log on to the computer with their LSU credentials; however, such logins do not count against their allotment of "5 LSU PC registrations".

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    SmartCheck

    SmartCheck [Entry in Progress - 01Feb2023] Advance Reservations, made through the Biosci-Booked (Genomics) [in progress], are required for use of this instrument (SmartCheck.jpg) in the Genomics Core. The SmartCheck provides a fast, practical way to verify that a pipette is dispensing accurately (±5% at 10 µl, 20 µl, 50 µl, 100 µl, 200 µl, 300 µl and 1000 µl). Simply dispense distilled water into the opening and the test volume will be automatically detected. Repeat this three times to obtain a Pass/Fail result.

    Thus, you can both quickly verify your pipette's performance immediately prior to 'volume-critical' experiments and know when you really need to have your pipettes re-calibrated. The SmartCheck can also be used to train (or re-train!) personnel in the proper operation of pipettes.

    You may also request that the Genomics Core check the performance of your pipettes on the SmartCheck platform (see GC Services, SmartCheck [in progress]).

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    QuantStudio·6 (qPCR)

    ABI QuantStudio-6, qPCR

    Advance Reservations, made through the Biosci-Booked (Genomics) instrument reservation system, are required for use of this instrument.

     

     

     

  • Submission of samples: It is critical that qPCR samples be submitted in accordance with the instructions presented below and at qPCR Samples.

  • Choice of qPCR plates and seals: Our qPCR platform is calibrated with ABI MicroAmp optical 96-well reaction plates and optical seals. Thus, the Background Calibration and the Dye Calibrations might not be correct for other brands of 'plastics', as they might not have the same fluorescent properties and might be more or less transparent to emissions from the instrument's thermal block. An extreme example is white plates which shield the camera from thermal block emissions, thereby switching the Background Calibration from being 'error-correcting' to 'error-inducing'.

    Thus, if you choose to use non-ABI 'plastics', best practice is to first run at least one test plate in which all 96-wells contain just nuclease-free water. This is not the same as having actually calibrated the qPCR platform with your choice of 'plastics', but it should give you a good idea as to whether your 'plastics' are at least compatible with the instrument's calibrations.

  • Create or analyze qPCR run files: Download v1.3 of QuantStudio-6 qPCR freeware (Windows only; no Mac version) [Warning: v1.7 is not compatible with our qPCR platform].
    – Upon first use of software, click 'Tools', 'Preferences', & 'Default'; select default instrument and 96-well standard block... and click 'Ok'.
    – On the 'Default' tab, you can also click the box for 'Show optical filters for run method'; this will add a second tab ('Optical Filters') to the Run Method screen under Experiment Setup. However, in most cases, you should not change the filters that are selected by default for your experiment type as doing so will limit your ability to correct Setup errors following completion of the run.
    – Submit run files (*.eds) by email, Genomics Core; completed run files will be returned to you by email.

  • Cradle-to-Grave” plate protection: All qPCR plates must be protected from picking up contaminates (on the outside of the plate) that might either fluoresce or quench fluorescence... from the time they are removed from the original manufacturer bag until they are placed into the block of the qPCR machine.

    Such contamination can be transferred to the block of the qPCR machine, adversely affecting your experiment and subsequent experiments until the block contamination is discovered and laboriously removed.

    Thus, your new plate must immediately go into a MicroAmp Splash-Free Support Base (a.k.a., rack; ThermoFisher Cat. No. 4312063), which is designed to minimize potential contact between the plate wells and the rack surface... unlike most racks which fit tightly around the plate wells. These qPCR racks.jpg are available from the Core as a "Supply".

  • Cold-Reaction Setup: In many cases, it is no longer necessary to set up qPCR under "cold conditions". However, if you do so, please note that ice may be contaminated even when visibly clean (as is evident upon melting a bucketful from typical ice machines).

    Thus, for "cold-reaction" setup, best practice is to use a dedicated metal block (chilled) for 96-well PCR-plates or a dedicated insulated container filled with clean ceramic or glass beads. If reactions are set up on ice, the plate must be centrifuged either immediately after being removed from the ice or after being briefly soaked in clean water.

    Finally, prolonged setup times with a chilled plate can result in atmospheric water condensing in the wells, thereby altering reaction volumes and ultimately your results. This problem can be severe under high humidity conditions (e.g., building A/C is not at normal capacity). One option is to cover the plate with a "cap" whenever not actively pipetting into the plate.

  • Applying Optical Adhesive Seals: Applying optical seals is not difficult; however, improper sealing technique can lead to evaporation from wells. For a primer on this topic, review ThermoFisher's YouTube video for proper Sealing Technique. (Caveat: Although the video voice-over states "between each well and along outside edges", please note that the demonstrator does not go "between each well" — which could disrupt well seals).

    It is also important to avoid touching the surface of the adhesive film as oils and other substances could affect its optical qualities or even fluoresce. Similarly, your applicator paddle must be clean. However, ethanol has its own fluorescent properties; thus, if you remove potential oils with ethanol, you need to thoroughly rinse the paddle with purified water and dry it with a Kimwipe.

    Finally, it is critical that you do not leave any sticky residue on the plate after removing the 'wings' from the adhesive film. Such residue can cause the plate to bond with the heated cover plate, making it difficult for the instrument to disengage from the plate.

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    Veriti thermal cycler

    Veriti thermocycler Advance Reservations, made through the Biosci-Booked (Genomics) instrument reservation system, are required for use of the Veriti thermocycler (located to the right of the qPCR platforms). This thermocycler performs standard PCR, including temperature gradients, and it is protected from brief power outages by an APC backup power supply. Normally, all other thermocyclers in the Genomics Core are reserved solely for use by the Core or members of the Batzer lab; however, you may request permission to use the other thermocyclers if you can plead extenuating circumstances.

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    Qubit 4.0

    Qubit_small
    If using your own assay kits, this fluorometer (Qubit-4.0.jpg) is available for use directly by you through the Biosci-Booked (Genomics) reservation system. Otherwise, samples are processed by the Genomics Core as described at the "Services" link below.

    There are many Qubit assays available from ThermoFisher. Assays require 1-20 ul of sample, resulting in the quantitation ranges shown with each kit. Samples are analyzed using Axygen 0.5-ml tubes (PCR-05-C, thin-wall, clear).

  • RNA IQ assay (integrity & quality; see description below)
  • dsDNA HS Assay Kit (0.2-100 ng)
  • dsDNA BR Assay Kit (2-1000 ng)
  • ssDNA Assay Kit (1-200 ng)
  • RNA HS Assay Kit (5-100 ng)
  • RNA BR Assay Kit (20–1,000 ng)
  • RNA XR Assay Kit (0.02-8 µg)
  • microRNA Assay Kit (1-1000 ng)
  • Protein Assay Kit (0.25-5 µg)

  • RNA IQ assay: In combination with the Qubit 4.0, this assay accurately distinguishes intact from degraded RNA by using two unique dyes: one that binds to large, intact RNA, and another that selectively binds to small, degraded RNA. Results are provided as an RNA IQ#, similar in concept to the RIN metric provided by a Bioanalyzer RNA assay. (Note: This kit is not compatible with earlier versions of the Qubit.)

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  • Services using Qubit 4.0
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    Diagenode Bioruptor NGS

    Diagenode_Bioruptor_NGS Advance Reservations, made through the Biosci-Booked (Genomics) instrument reservation system, are required for use of this instrument. Further, prior to each use of the Bioruptor, clients must ask the Genomics Core staff to prepare the system for operation. Staff will also provide instructions for operation of the Bioruptor, guidance on sonication profiles, and empty the system after use.

    To retain the integrity of DNA, Diagenode’s Bioruptor® NGS uses a gentle method of sonication and a chilled water bath to generate indirect sonication waves. As a result, the Bioruptor® produces better and more consistent results than are achieved with harsher sonication methods. Up to 12 closed tubes can be sonicated in parallel and the continuous rotation of tubes allows even distribution of the energy for efficient sonication.

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    ThermoFisher SPD1010 SpeedVac

    SpeedVac SPD1010 Advance Reservations, made through the Biosci-Booked (Genomics) instrument reservation system, are required for use of this instrument. This ThermoFisher instrument (SPD1010 SpeedVac.jpg) efficiently processes large batches of DNA or RNA samples.

    Currently, we have rotors for microtubes (1.5-ml, or 0.2-ml with inserts) and for 96-well plates. Typical applications include concentrating DNA extractions and removing low concentrations of non-agressive organic solvents (methanol, ethanol, acetonitrile) from samples. Most samples can be processed by simply pressing the "Start" button; however, users may also adjust settings for vacuum pressure, temperature, and run time.

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    Typhoon 8600 Variable Mode Imager

    Typhoon Variable Mode Imager Advance Reservations, made through the Biosci-Booked (Genomics) instrument reservation system, are required for use of this instrument. The Typhoon™ 8600 Variable Mode Imager is primarily used for storage phosphor experiments, but it is also capable of fluorescence detection of gels and blots. For DNA, RNA, and protein samples, users can choose from storage phosphor autoradiography, direct green (532 nm) excited fluorescence, and direct red (633 nm) excited fluorescence. The Typhoon can scan storage phosphor screens (mounted or unmounted), gels and blots (≤ 35 × 43 cm); further, images can be manipulated with ImageQuant™ Image Analysis Software.

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    Eppendorf Centrifuge 5810R

    Eppendorf Centrifuge 5810R At this time, we do not require reservations for this instrument. The Eppendorf 5810R Centrifuge is a refrigerated unit (-9 to 40°C), capable of handling both large tubes and microtiter plates. Maximum speed is ~2800 rcf, and a soft acceleration- deceleration option is available. The Genomics Core has the A-4-62 rotor, which sports two types of adaptors: (A) microtiter plates or 96-well PCR plates; and, (B) 15-ml or 50-ml tubes (with correct inserts). Other rotors could be obtained if there is sufficient interest.

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    Victor·3 Multilabel Counter

    MultiLable Counter This instrument is owned by Dr. Stephen Hand; thus, prior to your first use of the Victor·3, you must contact members of his lab (Rm. A627 & A629) for authorization and training. The Victor·3 Multilabel Counter (model 1420-011) includes fluorescence intensity (both top and bottom reading), luminescence, and absorbance (VIS) technologies as well as scanning, shaking and kinetics features. The instrument accepts all types of microtitration plates (from 1 to 1536 wells), as well as Petri dishes, slides, filters and Terasaki plates. Not all features described in the associated Victor·3 documents are applicable to this model. To learn how to operate the Victor·3, you can burn a CD of the software (available from the Genomics Core)and install it in “Demo Mode” on your personal computer.

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