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MJ Research DNA Engine Dyad PTC-220 Operation Manual

Peltier thermal cycler
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PTC-220 DNA Engine Dyad
™
Peltier Thermal Cycler
Operations Manual
Version 1.3
Boston • San Francisco • Tahoe • Copenhagen • Seoul
i

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Summary of Contents for MJ Research DNA Engine Dyad PTC-220

  • Page 1 PTC-220 DNA Engine Dyad ™ Peltier Thermal Cycler Operations Manual Version 1.3 Boston • San Francisco • Tahoe • Copenhagen • Seoul...
  • Page 2 PCR, and different rules may apply in different countries. Anyone who intends to use MJ Research equipment to do PCR is encouraged to contact Hoffmann-La Roche for more information, at one of the addresses below:...

  • Page 3: Table Of Contents

    Contents Documentation Conventions ............... iv Part I: The DNA Engine Dyad Thermal Cycler Introduction ..................1-1 Layout and Specifications ..............2-1 Installation ..................3-1 Operation ..................4-1 Creating Programs ................5-1 Managing and Editing Programs ............6-1 Running Programs ................7-1 Using the Utilities ................

  • Page 4: Documentation Conventions

    Documentation Conventions Before describing the various features of the Dyad cycler, let’s define some “common ground” conventions. • << >> will be used to indicate actual keys on the control panel, such as <<ENTER>>, <<1>> and <<LEFT>>. • < > will be used to indicate windowed menu items or buttons, such as <PROGRAMS>, <RUN>...

  • Page 5: Introduction

    Introduction Meet the DNA Engine Dyad Cycler, 1-2 Using This Manual, 1-2 Important Safety Information, 1-3...
  • Page 6 DNA Engine Dyad Operations Manual Meet the DNA Engine Dyad Cycler Thank you for purchasing an MJ Research DNA Engine Dyad™ thermal cycler. Designed by a team of molecular biologists and engineers, the DNA Engine Dyad cycler delivers multi- block thermal cycling with superior thermal performance. The programmable Dyad™ cycler, with its dual-bay chassis, is ideal for running multiple protocols and accommodating multiple users.
  • Page 7 Introduction Important Safety Information Safe operation of the DNA Engine Dyad cycler begins with a complete understanding of how the instrument works. Please read this entire manual before attempting to operate the Dyad cycler. Do not allow anyone who has not read this manual to operate the instrument. Warning: The DNA Engine Dyad cycler can generate enough heat to inflict serious burns and can deliver strong electrical shocks if not used according to the...
  • Page 8 DNA Engine Dyad Operations Manual...

  • Page 9: Layout And Specifications

    Layout and Specifications Front View, 2-2 Control Panel, 2-2 Back View, 2-3 Bottom View, 2-3 Alpha Units, 2-4 Single-Block Models, 2-4 Dual-Block Models, 2-4 Slide Block, 2-4 Power Bonnet Accessory, 2-4 Specifications, 2-5 Gradient Specifications, 2-5...
  • Page 10 DNA Engine Dyad Operations Manual Front View Dual-block Alpha (Figure 2-1) unit, lid closed Air exhaust vents Air intake vents Display screen Numeric keypad Touch pad Control Panel (Figure 2-2) M J R e s e a r c h E N T E R D N A E n g i n e P e l t i e r T h e r m a l C y c l e r...
  • Page 11 Layout and Specifications Back View Alpha unit handles (Figure 2-3) Power switch Ethernet port RS-232 port Power cord jack Fuses Bottom View (Figure 2-4) Dyad cycler front External mouse/touch pad Screen contrast adjustment knob External mouse port selection switch Air intake vents Dyad cycler rear...
  • Page 12 DNA Engine Dyad Operations Manual Alpha Units ™ Single-Block Models 60 Single: Holds 60 x 0.5ml tubes 96 Single: Holds 96 x 0.2ml tubes or one 96-well microplate 384 Single: Holds one 384-well microplate Flat Block: Holds customer-designed adapter through four screw- down points Dual-Block Models 30/30 Dual:...
  • Page 13 Layout and Specifications Specifications Thermal range: –5.0° to 105°C, but no more than 30°C below ambient temperature (4°C to 105°C, but not more than 23°C below ambient temperature for the Twin Towers unit) ® Accuracy: + 0.3°C of programmed target @ 90°C, NIST-traceable Thermal uniformity: + 0.4°C well-to-well within 30 seconds of arrival at 90°C (for most Alpha units;...
  • Page 14 DNA Engine Dyad Operations Manual...

  • Page 15: Installation

    Installation Packing Checklist, 3-2 Setting Up the DNA Engine Dyad Cycler, 3-2 Optional External Mouse Device, 3-2 Environmental Requirements, 3-3 Power Supply Requirements, 3-3 Air Supply Requirements, 3-4 Ensuring an Adequate Air Supply, 3-4 Ensuring That Air Is Cool Enough, 3-4 Requirements for Robotics Installations, 3-5 384-Well Microplate Specifics, 3-6...
  • Page 16 • Extended warranty application (US & Canada only) If any of these components are missing or damaged, contact MJ Research or the authorized distributor from whom you purchased the DNA Engine Dyad cycler to obtain a replacement. Please save the original packing materials in case you need to return the Dyad cycler for service.
  • Page 17 Note: Do not cut the supplied power cord and attach a different connector. Use a one-piece molded connector. If required, additional dedicated power cords may be purchased through MJ Research or authorized distributors.
  • Page 18 DNA Engine Dyad Operations Manual Air Supply Requirements The DNA Engine Dyad cycler requires a constant supply of air that is 31˚C or cooler in order to remove heat from the Alpha unit’s heat sink. Air is taken in from vents at the bottom and sides of the machine and exhausted from vents on both sides (see figures 2-1, 2-3, and 2-4).
  • Page 19 Installation Table 3-1 Troubleshooting Air Supply Problems l i t . s t Requirements for Robotics Installations Robotics installations require special attention to airflow and air temperature. Typically in these installations, DNA Engine Dyad cyclers and other thermal cyclers are restricted to a small area, along with other heat-generating equipment.
  • Page 20 DNA Engine Dyad Operations Manual 384-Well Microplate Specifics Some users find that a 384-well microplate can be difficult to remove from the 384-well block after completing their thermal cycling protocol. The plate fits very snugly in the block, and the 384 points of contact can provide a significant amount of friction. Fortunately, it is relatively simple to ameliorate this problem if it occurs in your application.
  • Page 21 Operation Turning the DNA Engine Dyad Cycler On, 4-2 Using the Control Panel, 4-3 Display Screen, 4-3 Operation Keys, 4-3 Block Status Lights, 4-4 Using the Data Ports, 4-4 Operating Alpha Units, 4-4 Installing an Alpha Unit, 4-4 Removing an Alpha Unit, 4-5 Opening an Alpha Unit, 4-6 Closing an Alpha Unit, 4-6 Selecting the Correct Sample Vessel, 4-6...
  • Page 22 DNA Engine Dyad Operations Manual Turning the DNA Engine Dyad Cycler On Caution: Do not insert or remove an Alpha unit with the DNA Engine Dyad cycler turned on; electrical arcing can result. Read the safety warnings in Ap- pendix A regarding electrical safety before inserting or removing an Al- pha unit or operating the Dyad cycler.

  • Page 23: Operation

    Operation Using the Control Panel The control panel (see figure 2-2) includes: a display screen, cursor keys, a numeric keypad with enter key, and a touch pad with left and right “mouse” buttons. Display Screen • The display screen is a 1/4 size VGA screen for displaying thermal cycler conditions and programs.
  • Page 24 DNA Engine Dyad Operations Manual Block Status Lights • When illuminated, these blue lights indicate whether the left and/or right Alpha units are in use. Using the Data Ports The DNA Engine Dyad cycler has two data ports located at the rear of the machine: an RS- 232 port and an Ethernet port.
  • Page 25 Operation Removing an Alpha Unit Caution: Do not insert or remove an Alpha unit with the DNA Engine Dyad cycler turned on; electrical arcing can result. Read the safety warning in Appen- dix A regarding electrical safety when inserting or removing an Alpha unit.
  • Page 26 Protocols may require some adjustment to ensure optimum performance when using a new vessel type. MJ Research offers a full range of tubes and microplates, manufactured to the specifications of each type of Alpha unit to ensure a precise fit. See Appendix 4-A of this chapter for a complete list.
  • Page 27 0.5ml tubes specifically designed for thermal cycling. The higher quality brands provide a good and consistent fit. MJ Research thin-walled 0.5ml tubes are designed for precise block fit and tight sealing of reactions down to 10µl.
  • Page 28 (–15° to –20°C). The samples are usually pure enough for analysis without an extraction. Chill-out liquid wax (available from MJ Research) is an easy-to-use alternative to oil. This purified paraffinic oil solidifies at 14°C and is liquid at room temperature. By programming a hold at low temperature, the wax can be solidified at the end of a run.
  • Page 29 Operation Microseal ® 'B' adhesive seals feature an aggressive adhesive, effective from –20°C to 110°C, which allows secure sample storage or transport before and after cycling. The clear polyes- ter backing allows easy inspection of sample wells. Microseal 'B' clear, adhesive seals are ideal for thermal cycling in all polypropylene and polystyrene microplates.
  • Page 30 Using Oil to Thermally Couple Sample Vessels to the Block With two exceptions (see below), MJ Research does not recommend using oil to thermally couple sample vessels to the block, for the following reasons: • Calculated-control protocols do not run accurately when oil is used.
  • Page 31 Operation Using the Optional Probe The probe consists of a precision thermistor mounted in a thin-walled plastic tube. A thin wire, encased in a small plastic tube, runs from the thermistor to the probe’s plug, which is inserted into a slot at the back of the Alpha unit (see figure 4-3). A small amount of oil is added to the probe tube to serve as the representative sample.
  • Page 32 Viscous oils are the best choice for the probe tube’s representative sample. They closely mimic the thermal characteristics of buffer solution, which changes temperature sluggishly due to the high specific heat of water. MJ Research recommends using heavy mineral oil, for the following reasons: •...
  • Page 33 Operation Detecting a Faulty Probe If the DNA Engine Dyad cycler detects that the probe is broken or missing when a protocol begins running, the protocol’s temperature control method is automatically switched from probe control to calculated control, and the following message is immediately displayed: "Probe Sensor Failure, Used Calc Control".
  • Page 34 System Selection Chart The following sample vessels and sealing options are recommended for use with the DNA Engine Dyad cycler and are available from MJ Research. To place an order, call 888-729-2165 or fax 888-729-2166. ✔ Reaction vessel fits block/sealing option fits reaction vessel without modification.
  • Page 35 Operation Appendix 4-B Safety Warning Regarding Use Of S Nucleotides Some researchers have experienced a problem with radioactive contamination when using S in thermal cyclers. This problem has occurred with all types of reaction vessels. The Problem When S nucleotides are thermally cycled, a volatile chemical breakdown product forms, probably SO .
  • Page 36 DNA Engine Dyad Operations Manual 2. If S must be used, three things will help control contamination: an oil overlay inside the tubes, mineral oil in the thermal cycler outside the tubes, and use of thick-walled 0.5ml tubes. Always run S thermal cycling reactions in a fume hood, and be aware that vessels may be contaminated on the outside after thermal cycling.
  • Page 37 Operation Appendix 4-C Using Silicone Oil in the Probe Tube The following light and heavy silicone oils may be used instead of mineral oil as the repre- sentative sample in a probe tube: • Dow Corning #200 light silicone oil (dimethypolysiloxane, Sigma #DMPS-5X) Density: 0.97g/ml Viscosity: 50cs Volume to use: 1.7 x volume of buffer in individual sample tube, plus one volume of oil...
  • Page 38 DNA Engine Dyad Operations Manual 4-18...

  • Page 39: Creating Programs

    Creating Programs Front Panel Setup, 5-2 Entering Program Steps, 5-14 Display Screen, 5-2 The Status Window, 5-14 Cursor Keys, 5-2 Entering a Program Using Graphical Mode, 5-15 Numeric Keypad, 5-3 Using the Mode Selection Window, 5-16 Touch Pad, 5-3 Using the File Save As Window, 5-17 Touch Pad Buttons, 5-3 The Graphical Programming Window, 5-19 Programming Conventions, 5-4...
  • Page 40 DNA Engine Dyad Operations Manual In this chapter, we will revisit the setup of the front panel, specifically those items used in program input. We will describe the conventions used, as well as the various programming steps and what they accomplish. We make suggestions regarding the translation of a cycle sequencing protocol into a Dyad program.
  • Page 41 Creating Programs Numeric Keypad This is located to the right of the cursor buttons and consists of a typical numeric keypad (numbers 0 through 9) and <<ENTER>> key. There is also a backspace/delete key and a decimal button. The numeric keypad is used to enter parameters such as temperature, hold time, and cycle iterations.
  • Page 42 DNA Engine Dyad Operations Manual Programming Conventions Before starting the Dyad programming process, let’s review some “common ground” conven- tions used here. • << >> will be used to indicate actual keys on the control panel, such as <<ENTER>>, <<1>> and <<LEFT>>. •...
  • Page 43 Creating Programs Temperature step: This sets incubation temperature and duration. The Dyad cy- cler ramps the sample to this temperature at its maximum rate unless ramp modifying instructions are added to the program (advanced mode only). The maximum rate of heating is 3°C/sec and cooling is 2°C/sec for all standard Alpha units (maximum rate of heating is 1.2°C/sec for the Twin Towers Alpha unit).
  • Page 44 DNA Engine Dyad Operations Manual Types of Programs There are two types of Dyad programs, graphical and advanced. Graphical programming features a graphical interface and a graphical representation of the program steps. Ad- vanced programming features a text-based interface and a descriptive listing of the program steps.
  • Page 45 Creating Programs Advanced Programs Advanced programs offer all of the Dyad programming features with the exception of the graphical programming interface. Creating an advanced program is desirable if you wish to enter a program that: • Contains many steps • Contains temperature or gradient steps with modifying instructions (i.e., increment temp, extend time, slow ramp, beep) •...
  • Page 46 Assume you have the necessary components for a 30-cycle sequencing reaction, and you have calculated the annealing temperature of your oligonucleotide to be 60°C. Please note that MJ Research recommends using 92°C as the default denaturation temperature during cycling steps. The resulting raw program you write may look something like this: Raw program: 92°C for 30 seconds...
  • Page 47 Creating Programs Considerations During Program Creation Once you have written the body of your raw program, there are decisions to make before creating your Dyad program. They concern how your steps should be implemented. These decisions involve the following: • Temperature control mode •...
  • Page 48 The Dyad cycler adjusts the block’s temperature to maintain the probe at programmed temperatures. You may purchase an in-sample probe from MJ Research, Inc. The in- sample probe is comprised of a temperature probe placed inside a typical capped tube.
  • Page 49 Creating Programs Modifying a Program Designed for a Different Machine The ramp programming step can be used to adapt programs designed for thermal cyclers with slower maximum heating and cooling rates than the Dyad cycler. In addition, a given protocol will occasionally work better with a slower rate of temperature change; the ramp step can be used to optimize the program for such a protocol.
  • Page 50 DNA Engine Dyad Operations Manual Choosing a Temperature Hold Time Because of the calculated melting temperature (Tm) of a DNA hybrid, DNA polymerase processivity, and reaction kinetics, it may be possible to generalize conditions regarding thermal-cycling protocols. However, decisions on denaturation, annealing or extension hold times will be reaction specific and should be optimized.
  • Page 51 Creating Programs Your written program should now appear as follows: Raw program: Use calculated temperature control mode Use constant lid control mode at 100°C 92°C for 30 seconds Gradient from 45°C to 65°C for 3 minutes Goto step 1, 29 more times Beyond the Example Protocol: Other Considerations In addition to the above considerations, you can also include other protocol variations which...
  • Page 52 DNA Engine Dyad Operations Manual Entering Program Steps From our example, we are ready to enter a new program. When executing a selected command via the touch pad, we will use the method of tapping the pad with a fingertip. This can also be accomplished with the left button below the touch pad, as well as the <<EN- TER>>...
  • Page 53 Creating Programs The Program Display box will list steps for the program currently running on the selected block. The menu bar at the top of the Status window includes five submenus: <Programs>, <Com- mand>, <Tools>, <View>, and <Utilities>. These submenus provide the operator with paths for maneuvering through the various Dyad software windows.
  • Page 54 DNA Engine Dyad Operations Manual Begin by choosing the temperature control mode and lid control mode for the program. Refer to the “Considerations During Program Creation” section earlier in this chapter for information on temperature and lid control modes. The current mode of temperature control is listed in the Control Mode field.
  • Page 55 Creating Programs Place the cursor in the Maintain lid temperature at field and select the field. We have decided to set the lid to a constant temperature of 100°C. • Enter 100 from the numeric keypad. Place the cursor in the below field and select the field. We have decided to turn the lid off when the block drops below 30°C.
  • Page 56 DNA Engine Dyad Operations Manual The File Save As window presents the operator with a space for entering the program name. The program will be added to the folder indicated in the Folder field. The <New Folder> button creates a new folder in which to store your new program.
  • Page 57 Creating Programs You will be returned to the File Save As window. The display will have changed slightly, with our newly created folder appearing in the Folder list. We will want to save our program in this newly created folder. •...
  • Page 58 DNA Engine Dyad Operations Manual Selecting a Step When a step is selected, the line(s) or arrow depicting that step will turn red. There are several ways to select a step. You can select a step by positioning the cursor in the step’s column and tapping or clicking once.
  • Page 59 Creating Programs Entering a Temperature Step Recall again our raw program: Use calculated temperature control mode Use constant lid control mode at 100°C An initial incubation at 94°C for 1 minute 92°C for 30 seconds Gradient from 45°C to 65°C for 3 minutes Goto step 1, 29 more times An incubation at 10°C forever The first actual step in the protocol is the incubation at 94°C for 1 minute.
  • Page 60 DNA Engine Dyad Operations Manual 3. If the first step displayed is a temperature step, or if a temperature step was added as indicated in scenarios 1 or 2: • Position the cursor (text pointer) in the temperature field of step 1 and select by tapping/clicking once.
  • Page 61 Creating Programs Step one of our protocol now consists of a temperature step with an incubation tempera- ture of 94.0 and a duration of 01:00. Step 1 temperature step Recall that the maximum programmable temperature is 105.0°C and the minimum pro- grammable temperature is 0.0°C in a graphical program.
  • Page 62 DNA Engine Dyad Operations Manual Step two of our protocol should now specify a temperature incubation of 92.0 for a duration of 00:30. Step 2 temperature step Entering a Gradient Step Step three of our program is a gradient step designed to determine the optimal anneal- ing temperature of our oligonucleotide.
  • Page 63 Creating Programs • Tap or click once to accept the changes to the gradient range. Note: Change both the higher and lower temperatures before accepting the changes to the gradient step to ensure that the temperature differential is not greater than 24°C or less than 1°C. •...
  • Page 64 (e.g., 4°C)—but, colder temperatures require considerably more power to maintain, and are unnecessary in most circumstances, in the opinion of MJ Research scientists. Adding a forever incubation step is identical to programming a temperature step with the exception of selecting the <Forever>...
  • Page 65 Creating Programs Unused temperature step Step 5 forever temperature step If there are any additional steps displayed in the graphical programming window that you do not wish to include in the program, select the step, and then select <Delete Step>. We do not wish to include the temperature step displayed in the fifth temperature/gradi- ent step column.
  • Page 66 DNA Engine Dyad Operations Manual Entering a Program Using Advanced Mode • Select <Programs>. Note: As described earlier, this involves positioning the screen cursor over <Pro- grams> with a fingertip on the touch pad and tapping the touch pad once. Drop-down submenus appear, including <Open>, <New>, <Copy>, <Move>, <Delete>, <Delete Folder>, and <New Folder>.
  • Page 67 Creating Programs It is from the advanced programming window that you will add steps using the <Temp>, <Gradient>, <Goto>, and <Lid> options. Additionally, buttons running across the window bottom provide options to <Delete Step>, <Save + Run>, <Save>, or <Save As> programs, and <Cancel> the current programming ses- sion.
  • Page 68 DNA Engine Dyad Operations Manual Entering a Temperature Step While the program ADV#1 is a bona fide Dyad program, it has no utility. A run of this program will finish immediately after its start, because there are no temperature com- mands or incubation times to constitute an actual run.
  • Page 69 Creating Programs The Temperature Step window presents the operator with a number of tempera- ture adjustment options. The Temperature and Time fields as well as the Beep on Target selection option are available at the top of the window. At the bottom, the Increment Temp , Extend Time and Slow Ramp options are available, each with their own <Set Parameters>...
  • Page 70 DNA Engine Dyad Operations Manual • Select <Temp>. • Position the cursor over the Temperature field and select. • Enter 92 from the numeric keypad into the field. • Similarly, enter 30 in the Time:Sec field. Additionally, we want the cycler to beep after reaching the target temperature. •...
  • Page 71 Creating Programs Entering a Gradient Step The Gradient Step window indicates the step number and includes fields for specifying the Lower Temperature and the Higher Temperature of the gradient range as well as the gradient hold Time . The maximum gradient range is 24°C and the minimum range is 1°C.
  • Page 72 DNA Engine Dyad Operations Manual The newly created gradient can be previewed graphically. • Select <Preview>. This distribution of temperatures specified by the gradient should be reviewed and any changes to the gradient limits made before a program run. Please note that the gradient temperature differential is not linear, with a broader spread in temperature between the center columns of wells.
  • Page 73 Creating Programs As discussed previously, increasing the time per cycle of a temperature step might be desirable in some protocols where extra time is required during later cycles to allow synthesis to be completed. We will not require such a step in our cycle sequencing protocol, but describe it here for completeness.
  • Page 74 DNA Engine Dyad Operations Manual • Enter 5 in the field. • Select <OK>. This program step, incorporating a per cycle increase in incubation time, would appear as follows: Incubate at 60°C for 00:01:00 Increase by 5.0 seconds every cycle This step is not included in our cycle sequencing protocol, so we continue with our next addition.
  • Page 75 Creating Programs The program now appears as follows: Temperature Control Mode: Calculated Lid Control Mode: Constant at 100°C 1. Incubate at 94°C for 00:01:00 Beep on Target 2. Incubate at 92°C for 00:00:30 Beep on Target 3. Gradient from 45°C to 65°C for 00:03:00 4.
  • Page 76 DNA Engine Dyad Operations Manual ° • Position the cursor over the C per second field and select. • Enter 0.5. • Select <OK>. You would expect the program step as included in a protocol to appear as fol- lows: Incubate at 60.0°C for 00:00:30 Ramp to 60°C at 0.5°C per second The Increment Temp option...
  • Page 77 Creating Programs • Select Decrease . ° • Position the cursor over the by __ C per cycle field and select. • Enter 0.2 in the field. • Select <OK>. You would expect the program step as included in a protocol to appear as fol- lows: Incubate at 60.0°C for 00:00:30 Decrease by 0.2°C every cycle...
  • Page 78 DNA Engine Dyad Operations Manual 5-40...

  • Page 79: Managing And Editing Programs

    Managing and Editing Programs Opening a Program, 6-2 Opening a Program in Advanced Mode, 6-2 Opening a Program in Basic Mode, 6-4 Editing a Program, 6-4 Editing a Graphical Program, 6-5 Editing an Advanced Program, 6-6 Highlighting and Selecting Program Steps, 6-6 Deleting a Step, 6-6 Inserting a Step, 6-7 Editing a Step, 6-7...
  • Page 80 DNA Engine Dyad Operations Manual In the previous chapter, various entering and editing features were discussed as they applied to entering a graphical and/or an advanced program. In this chapter, we cover in more depth the options available for the manipulation of existing Dyad programs. The programming conventions listed in Chapter 5 will also be used here.
  • Page 81 Managing and Editing Programs We had previously saved ADV#1 in the FOLDER2 folder. • Select the folder FOLDER2. • Select the program ADV#1. • Select <OK>. You are presented again with the advanced programming window. It is from this window that steps can be inserted, deleted, or edited. While all graphical programs can be opened and edited in advanced mode, only a subset of advanced programs can be opened and edited in basic mode.
  • Page 82 DNA Engine Dyad Operations Manual Opening a Program in Basic Mode You can choose to open and edit a graphical program in either the graphical programming window, or in the advanced programming window. Opening a graphical program in ad- vanced mode is desirable if you wish to add step modification options, incubations below 0°C, or other programming features not available in graphical programs (see the “Types of Programs”...
  • Page 83 Managing and Editing Programs After running the above program, analysis of the resulting sequencing data indi- cated that 60°C was the best annealing temperature. We would like to change step 3 from a gradient step into a temperature step. Further, step 2 includes a 30 second incubation that we wish to change to 25 seconds.
  • Page 84 DNA Engine Dyad Operations Manual Editing an Advanced Program Highlighting and Selecting Program Steps Several important factors determine step selection: • The insertion point for new steps is AFTER the step highlighted in the program listing. This highlighting is accomplished with a SINGLE tap or click. •...
  • Page 85 Managing and Editing Programs Inserting a Step Now we will insert an annealing temperature step. • Highlight step 2. Recall that new steps are inserted AFTER the highlighted step. • Select <Temp> from the advanced programming window. The Temperature Step window appears. Select an incubation Temperature of 60°C, a hold Time of 3 minutes, and the Beep on Target option.
  • Page 86 DNA Engine Dyad Operations Manual Now the Program Editing window lists the program steps as the following: Temperature Control Mode: Calculated Lid Control Mode: Constant at 100°C 1. Incubate at 94°C for 00:01:00 Beep on Target 2. Incubate at 92°C for 00:00:25 Beep on Target 3.
  • Page 87 Managing and Editing Programs Copying a Program • Select <Programs> in the Status window. • Select <Copy> from the drop-down menu. Once you have created a number of programs, you may want to create separate folders to organize them. Perhaps you will use separate folders for different us- ers, or experimental series.
  • Page 88 DNA Engine Dyad Operations Manual Deleting a Program • Select <Programs> in the Status window. • Select <Delete> from the drop-down menu. Important!: Use caution when deleting programs, as deletions are irreversible, and the delete program window is very similar to the open program window. We provide a conve- nient program listing at the bottom of the window so that you can determine whether you truly wish to delete the program.
  • Page 89 Managing and Editing Programs Moving a Program • Select <Programs> in the Status window. • Select <Move> from the drop-down menu. Moving is the same as copying (described above), with one distinction: only one copy of the program is maintained in the To Folder . The copy in the From Folder is deleted. You will not be prompted with a verification step for this move, so exercise some caution.
  • Page 90 DNA Engine Dyad Operations Manual You will be presented with a confirmation screen asking you to verify the dele- tion. Select <Yes> to delete the folder. In this chapter, we have utilized editing tools to add and subtract steps from our example programs.
  • Page 91 Running Programs Using the Instant Incubation Feature, 7-2 Running Programs, 7-4 The Run Program window, 7-5 During the Run, 7-6 Run Status, 7-6 Terminating a Run, 7-7 Pausing/Resuming a Run, 7-7 Skipping a Step, 7-8 Inaccessible Features, 7-8 Running Multiple Programs, 7-8...
  • Page 92 DNA Engine Dyad Operations Manual In Chapter 5, we translated an experimental protocol into functional Dyad programs. In Chapter 6, we edited program steps. In this chapter, we explore the implementation of the Dyad programs, and the instant incubation feature. Using the Instant Incubation Feature The DNA Engine Dyad cycler can also be quite useful as an “instant”...

  • Page 93: Running Programs

    Running Programs • Select the Temperature field and enter the desired temperature. • Select Heated Lid if you are incubating at a high temperature and wish to minimize condensation (refer to the “Sealing with the Hot Bonnet Lid” section in Chapter 4 for additional information on us- ing the heated lid).
  • Page 94 DNA Engine Dyad Operations Manual Running Programs Note: The Operations Chapter (Chapter 4) provides information on running programs with respect to the operation and use of Alpha units. For example, a program containing a gradient step will only run in 96-well Alpha units. Please review this chapter for additional technical detail, to ensure that the program you load and run is an appropriate match for your Alpha units.
  • Page 95 Running Programs You are now presented with the Run Program window. The Run Program window The Run Program window allows you to select the Block(s) that you wish to run your protocol on. You can select a single block or all blocks (based on block compatibility with your program).
  • Page 96 DNA Engine Dyad Operations Manual vessels used. • Select <OK> to initiate the run. Refer to the “Running Multiple Programs” section at the end of this chapter for information on simultaneously running multiple programs. During the Run Run Status Information in the Temperature , Time, and Cycle fields of the Status window will indicate that your program is running.
  • Page 97 Running Programs Tip: Recall that convenient and rapid toggling between the Status and Graphs windows can be achieved by selecting the button in the lower right of the Status and Graphs windows respectively. To simultaneously view the status of all blocks including the Block Name , Block Status , the name of the Program Running , the Time Remaining in the program, the Time Elapsed , and the User name, select the <View>...
  • Page 98 DNA Engine Dyad Operations Manual To resume a run, select <Resume> from the bottom of the Status window or select <Com- mand>, <Pause/Resume>, and either <Resume All> or select the desired block from the drop-down list. Skipping a Step To skip to the next step in a running program, select <Skip> at the bottom of the Status window.
  • Page 99 Running Programs To run multiple programs, first choose an available block from the block selection menu in the Status window. Available blocks will show a “Block is Inactive” message just above the User Name field. If there are no available blocks, the “No blocks available” message window appears.
  • Page 100 Using the Utilities <About>, 8-2 <User Name>, 8-3 <Set Date/Time>, 8-3 <Remote>, 8-4 <Auto Remote>, 8-4 <Update Soft> & <Network Config>, 8-5 <Ping>, 8-7 Additional Utilities, 8-8 Transferring Program Files, 8-8...
  • Page 101 DNA Engine Dyad Operations Manual In previous chapters, you’ve learned how to install and operate the Dyad cycler, as well as write and execute programs. In this chapter, the various functions of the <Utilities> submenu will be discussed. The <Utilities> submenu rounds out the capabilities of the Dyad cycler, providing access to user name control, remote command mode, and the date and time settings.

  • Page 102: Using The Utilities

    Using the Utilities <User Name> When this item is selected, the following screen will appear: By selecting <Add User Name>, the virtual keyboard will appear allowing users to enter their name prior to program execution. In this manner, any potential user can determine the “owner”...
  • Page 103 DNA Engine Dyad Operations Manual <Remote> To operate a Dyad cycler in remote mode (i.e., from a desktop computer), select <Remote> from the <Utilities> menu. The Remote Mode window will appear on the screen. The only option available is a check box for enabling syntax checking. This will filter out any commands that do not conform to the PTC Remote Command Set syntax.
  • Page 104 Using the Utilities <Update Soft> & <Network Config> These utilities allow you to update both the software controlling the Dyad cycler’s hardware (i.e., engine software), and the user-interface/programming software (i.e., front end soft- ware) by establishing an internet connection and directly downloading any new software upgrades or versions.
  • Page 105 DNA Engine Dyad Operations Manual If you have a static internet connection, i.e., the IP address does not change, select the Static option. Enter your IP Address , Net Mask , GateWay , and DNS Server information using the numeric keypad. If you lack this information, please contact your network administrator. If you have a dynamic internet connection, i.e., a different IP address is assigned each time you access the internet, select the DHCP (Dynamic Host Configuration Protocol) option.
  • Page 106 Using the Utilities <Ping> This utility can be used to verify that the ethernet software and connection are properly functioning. Select <Ping> from the <Utilities> menu. In the Ping window, begin by selecting <Host Names> and entering the IP address of the computer that you wish to ping.
  • Page 107 DNA Engine Dyad Operations Manual Minimize button Close button (select to exit) (do not select) Select <Ping!> to repeat the ping, or select the minimize button to exit the window. Note: Do not exit this window by using the close button. If you inadvertently exit using the close button, an error will appear directing you to power cycle the instrument.
  • Page 108 Using the Utilities 4. To execute the file transfer, select Proceed (& Exit when completed) by entering 1 on the numeric keypad of the sending instrument. Once the transfer is complete, the TRANSFER COMPLETE message will be displayed and the instrument will proceed with the boot sequence.
  • Page 109 DNA Engine Dyad Operations Manual 8-10...

  • Page 110: Maintenance

    Maintenance Cleaning the DNA Engine Dyad Cycler, 9-2 Cleaning the Chassis and Block, 9-2 Cleaning the Air Vents, 9-2 Cleaning Radioactive or Biohazardous Materials Out of the Block, 9-3 Changing the Fuses, 9-3...
  • Page 111 If you use oil in the block (a practice not recommended by MJ Research, Inc.; see “Using Oil to Thermally Couple Sample Vessels to the Block,” in Chapter 4), clean the wells whenever the oil has become discolored or contains particulate matter.
  • Page 112 Maintenance ✔ Tip: To prevent problems with overheating, institute a regular program of checking for dust buildup, particularly for robotics installations. Cleaning Radioactive or Biohazardous Materials From the Block When cleaning machines that have been running radioactive or biohazardous reactions, consult your institution’s radiation safety officer or biosafety officer regarding cleaning meth- ods, monitoring, and disposing of contaminated materials.
  • Page 113 DNA Engine Dyad Operations Manual...

  • Page 114: Troubleshooting

    Troubleshooting Sources of Problems, 10-2 System Problems, 10-2 Error Messages, 10-3 Problems in Power-Up, 10-8 Problems with System Performance, 10-9 Problems with an Alpha Unit, 10-10 Problems Related to Protocols, 10-10 Problems due to Environmental Conditions, Setup, and Maintenance, 10-13...

  • Page 115: Sources Of Problems

    • Problems related to protocols When troubleshooting a difficulty, it is advisable to have the following pieces of information available, should it become necessary to contact MJ Research, Inc. for assistance: • Serial and catalog numbers for the base as well as Alpha units •...

  • Page 116: Error Messages

    Troubleshooting Table 10-1: Error Messages The Error Codes listed in the table below correspond to the error codes returned in the Dyad cycler’s <Error Log>. The Remote Error Codes listed correspond to the error codes returned when the Dyad cycler is operated in remote mode (see the <Remote> and <Auto Remote> sections in Chapter 8 for more information on remote mode).
  • Page 117 DNA Engine Dyad Operations Manual Remote Error Code Reason Action Error Code Please observe the following guidelines: 1) place instruments at least 10cm apart, 2) avoid placing instruments in any location where hot air might The heat sink temperature is enter the intake vents, 3) place registering somewhat higher instrument on a hard surface with...
  • Page 118 Troubleshooting Remote Error Code Reason Action Error Code Call (888) MJ CYCLE (in the US The internal fan is not providing or Canada), or your local adequate cooling for the distributor (outside the US or instrument. Canada). An irregularity has been detected in a heated lid sensor.
  • Page 119 DNA Engine Dyad Operations Manual Remote Error Code Reason Action Error Code Please observe the following guidelines: 1) place instruments at least 10cm apart, 2) avoid placing instruments in any location where hot air might The power supply temperature enter the intake vents, 3) place was higher than normal.
  • Page 120 Troubleshooting Remote Error Code Reason Action Error Code A program containing a If you place a 96-well Alpha unit None gradient step can only be run in this quadrant, you may run on a 96-well Alpha unit. this gradient program. The program entered may only run in Calculated or Block temperature control mode.

  • Page 121: Problems In Power-Up

    DNA Engine Dyad Operations Manual Problems in Power-Up Should a Dyad cycler not power up properly, as indicated in Chapter 4, please follow the steps outlined in the flowchart (figure 10-1) to determine the best course of action. In this case, a problem should be consistent and repeatable.

  • Page 122: Problems With System Performance

    Troubleshooting Problems with System Performance Should you encounter problems with menu navigation, front panel manipulation, or any performance aspect of a Dyad cycler that has successfully powered up, please follow the steps in this flowchart to determine the best course of action. Again, a problem should be a consistent, repeatable problem.

  • Page 123: Problems With An Alpha Unit

    An Alpha unit is a distinct, separately engineered piece of equipment that is made to inter- face with all MJ Research DNA Engine line thermal cyclers. However, to rule out problems with any Alpha unit, please follow this recommended troubleshooting flowchart.
  • Page 124 Troubleshooting t s i i l i s ' l > ° 2 Only rarely are temps higher than 94°C required, however. t s i i l i h, Inc. l i f l l i . l i Test minus sample, with a control template and primer set.
  • Page 125 DNA Engine Dyad Operations Manual Table 10-2: Protocol Difficulty P " " r < . s l . s l p " " r - f l t i r p " " . s . s t t s i i l i s ' l t i r...

  • Page 126: Problems Due To Environmental Conditions, Setup, And Maintenance

    Troubleshooting Problems due to Environmental Conditions, Setup, and Maintenance Strict adherence to the installation, operation, and maintenance instructions provided in Chapters 3, 4, and 9 are tantamount to the continued trouble free operation of the Dyad cycler. Should it be determined that the source of a problem is due to incorrect operation or setup, consult Table 10-3 for a list of problems and suggested solutions.
  • Page 127 DNA Engine Dyad Operations Manual 10-14...

  • Page 128: Alpha Units And The Remote Alpha Dock System

    Alpha Units and the Remote Alpha Dock System Alpha™ Units Available from MJ Research, Inc., 11-2 About the Remote Alpha Dock System, 11-4 Packing checklist, 11-5 Requirements, 11-5 Environment, 11-5 Power Supply, 11-6 Air Supply, 11-7 Installation, 11-7 Operation, 11-8...
  • Page 129 The “384” for 384-well Plates (ALS-1238) MJ Research, Inc. has worked with several genome centers to develop a high-density format for automated operation. The result is the 384-well Alpha unit, a true 4X version of the 96- well format. The wells have the same V-profile and are spaced on 4.5mm centers.
  • Page 130 This sample format can hold one 384-well microplate and features a Power Bonnet motor- ized lid for remote control of lid opening and closing. Contact MJ Research, Inc. at (888) 735-8437 for additional information on the Alpha units listed here, as well as updates on any new Alpha units available.
  • Page 131 The Remote Alpha Dock™ system is designed to add flexibility to the installation and opera- tion of the MJ Research PTC-220 DNA Engine Dyad cycler. The system allows Alpha units to be placed at a distance from the PTC-220 base, enabling more efficient use of space and facilitating robotic operation.
  • Page 132 • Extended warranty application (US and Canada only) Requirements Environment The Remote Alpha Dock system allows for custom installations. The following placement configurations are recommended by MJ Research, Inc. Figure 11-2 Remote Alpha Dock base and dock connector, bottom view. 11-5...
  • Page 133 DNA Engine Dyad Operations Manual • Remote Alpha Dock bases with mounted Alpha units may be configured in any horizon- tal orientation or array as long as a minimum side clearance of 10 cm is maintained between the Remote Alpha Dock unit and any wall, bulkhead, or adjacent unit (this is identical to the PTC-220 base requirement).
  • Page 134 Alpha Units and the Remote Alpha Dock System Air Supply • Alpha units being operated in the remote configuration have no operating constraints that do not also apply to normal operations in the PTC-220. Installation • Turn the dock connector upside down, so that the green circuit board is visible. Check that the ground lead remains attached at both ends (fig.
  • Page 135 DNA Engine Dyad Operations Manual • Connect the multi-pin power cable’s female end to the connector labeled “Power Port” and slide the latch to lock the pins in place. • Attach the data cable’s “DOCK DATA” end to the connector labeled “Data Port” and slide the latch to lock the pins in place.
  • Page 136 Alpha Units and the Remote Alpha Dock System Figure 11-5 Flush-mounting template 11-9...
  • Page 137 DNA Engine Dyad Operations Manual 11-10...

  • Page 138: Appendix A: Safety Warnings And Guidelines

    Appendix A: Safety Warnings and Guidelines Appendix A Safety Warnings and Guidelines Warning: When removing an Alpha unit from a PTC-220 DNA Engine Dyad base, keep all fingers and foreign objects away from the Alpha unit bays. Keep all objects clear of the Alpha unit bays until the fan has come to rest. Warning: Operating the PTC-220 DNA Engine Dyad cycler before reading this manual can constitute a personal injury hazard.
  • Page 139 DNA Engine Dyad Systems Operations Manual Safe Use Guidelines The PTC-220 DNA Engine Dyad cycler is designed to be safe to operate under the following conditions: • Indoor use • Altitude up to 3000m • Ambient temperature 5°C–31°C • Relative humidity 10–90%, noncondensing •...

  • Page 140: Appendix B: How A Peltier Heat Pump Works

    Appendix B: How a Peltier Heat Pump W orks Appendix B How a Peltier Heat Pump Works The functional heart of every DNA Engine Dyad cycler is a high-performance Peltier-effect heat pump (also known as a “thermoelectric module”). The “MJ” module is a solid-state device manu- factured to withstand the thermal stresses associated with rapidly cycling temperatures.

  • Page 141: Appendix C: Shipping Instructions For Us Residents

    Users residing in the United States should follow these instructions for shipping a machine to MJ Research for factory repair or an upgrade. Users outside of the United States should send machines to their distributor, in accordance with shipping instructions obtained from the distributor.

  • Page 143: Appendix D: Warranties

    If a defect is discovered, MJ Research will, at its option, repair, replace, or refund the purchase price of the THERMAL CYCLER at no charge to the customer, provided the product is returned to MJ Research within the warranty period.
  • Page 144 The coverage must be purchased through a purchase order received by MJ Research within 30 days of receipt of the offer of extended warranty or the offer to renew the extended warranty. These offers apply only to machines...

  • Page 145: Index

    Index Index About utility 8-2 Accessories. See Power Bonnet lid ; Remote Alpha Dock system Accuracy. See Thermal accuracy Adding a step, advanced program 6-7 Adding a step, graphical program 5-20 , 6-5 Advanced mode, programming 5-28–5-39 Advanced program deleting a step 6-6 editing 6-6–6-8 .
  • Page 146 DNA Engine Dyad Operations Manual Basic mode, programming 5-15–5-39 Bleach, using in block 9-2 Block control 5-10 Block selection incompatibility 7-8 Block status lights 4-4 Calculated control 5-9 Chill-out liquid wax 4-8 Cleaning 9-2 air vents 9-2 biohazardous materials 9-3 chassis and block 4-16 , 9-2 radioactive materials 4-16 , 9-3 removing oil from block 9-2...
  • Page 147 Index Editing a program 6-4–6-8 advanced 6-6–6-8 deleting a step 6-6 editing a step 6-7–6-8 inserting a step 6-7 selecting steps 6-6 graphical 6-5–6-6 deleting a step 5-20 editing a step 5-20 , 6-5 inserting a step 5-20 , 6-5 selecting steps 5-20 Editing a step, advanced program 6-7–6-8 Editing a step, graphical program 5-20 , 6-5...
  • Page 148 DNA Engine Dyad Operations Manual Goto step 5-5 , 5-8 advanced program 5-36–5-37 graphical program 5-25–5-26 Gradient step 5-5 advanced program 5-33–5-35 graphical program 5-24–5-25 Gradient, using to optimize 5-12 Gradients gradient preview window 5-34 specifications 2-5 accuracy 2-5 calculator accuracy 2-5 column uniformity 2-5 lowest/highest temperature 2-5 temperature differential range 2-5...
  • Page 149 Index In sample probe control 5-10 Increment temp option 5-5 , 5-38–5-39 Inserting a step, advanced program 6-7 Inserting a step, graphical program 5-20 , 6-5 Instant incubation 7-2 Keyboard, virtual 5-18 Keypad 5-3 Layout back view 2-3 bottom view 2-3 control panel 2-2 front view 2-2 Lid control methods...
  • Page 150 DNA Engine Dyad Operations Manual Network config utility 8-5–8-6 Numeric keypad. See also Control panel location 2-2 Opening a program 6-2–6-4 advanced mode 6-2–6-3 basic mode 6-4 Operation turning the Dyad cycler on 4-2 Packing checklist 3-2 Password utility 8-8 Pausing a run 7-7–7-8 Peltier effect B-1 Ping utility 8-7–8-8...
  • Page 151 Index Program advanced 5-28–5-39 extend time option 5-35 goto step, entering 5-36–5-37 gradient step, entering 5-33–5-35 increment temp option 5-38–5-39 lid control step, entering 5-37 slow ramp option 5-37–5-38 temperature step, entering 5-30–5-33 advanced options beep on target 5-5 extend time 5-5 increment temp 5-5 slow ramp 5-5 copying 6-9...
  • Page 152 DNA Engine Dyad Operations Manual monitoring the run graph window 7-6 status window 7-7 moving 6-11 opening 6-2–6-4 advanced mode 6-2–6-3 basic mode 6-4 running 7-4 run program window 7-5 running multiple 7-8 saving 5-17–5-19 , 6-8 types 5-6–5-7 advanced 5-7 graphical 5-6 Programming conventions 5-4 Programs menu 6-8...
  • Page 153 Index Safety general instructions 1-3 guideline for safe use A-2 S Nucleotides 4-15 Use of warnings A-1 Sample vessels ensuring good thermal contact 4-10 loading into block 4-10 sealing reason for 4-8 selection chart 4-14 with Hot Bonnet lid and caps/film 4-8 with oil or wax 4-8 selection 4-6 0.2ml tubes 4-7...
  • Page 154 DNA Engine Dyad Operations Manual Temperature control methods block control 5-10 calculated control 5-9 choice of calculated control over probe control 4-11 in sample probe control 5-10 modifying a program from a different machine 5-11 modifying block and probe control programs 5-10 Temperature hold time 5-12 Temperature increment.
  • Page 155 Index service 8-8 set date/time 8-3 update soft 8-5–8-6 user name 8-3 Virtual keyboard 5-18 Warranties D-1 In-11...
  • Page 156 DNA Engine Dyad Operations Manual In-12...

  • Page 157: Declaration Of Conformity

    Declaration of Conformity DoC-1...

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