Fungal DNA Barcoding Primer Pairs (ITS1F/ITS4), 10 µM, 200 µLbeta
£16.59
Fungal DNA Barcoding Primer Pairs (ITS1F/ITS4), 10 µM, 200 µLbeta
£16.59
Use these primer pairs for PCR or sequencing purposes when DNA barcoding fungi.
The primers are pre-prepared at a standard working concentration (10 µM) in low TE buffer for easy use. For PCR, use in forward-reverse pairs and add to the PCR mix before adding DNA template. For Sanger sequencing, use as specified by your sequencing company. The 200 µL tube pairs are suitable for up to 500 PCRs of 20 µL (0.4 µL/rxn).
Primers are short DNA fragments of a specific sequence that are used to define the region amplified during PCR.
When used as part of a PCR, the primers ITS1F and ITS4 allow amplification of the Internal Transcribed Spacer (ITS) region: the most commonly used and useful single DNA barcode region for fungi. The ITS region contains the ITS1, 5.8s, and ITS2 regions.
The primers are formulated at a standard concentration (10 µM), and can be used as a pair for PCR, or sent individually to Sanger sequencing facilities for forward or reverse sequencing.
Details
Ready-To-Use: Skip rehydrating your primers, and just add to your PCR mix.
Versatile: Use as pairs for PCR, or individually for forward or reverse sequencing when sent to Sanger sequencing services. The individual primers can also be combined with other forward or reverse primers if needed.
Applications
For use in fungal DNA barcoding and Sanger sequencing.
Use these primer pairs as part of a PCR mix to amplify the internal Transcribed Spacer (ITS) DNA barcode region of fungi.
Use the primers individually for sending to Sanger sequencing services to act as a primer for the Cycle sequencing step of sequencing.
Specifications
Recommended Usage
Primers are typically used at between 0.1 µM to 1 µM final concentration. We recommend 0.2 µM for typical usage.
In a 20 µL PCR, use 0.4 µL of each primer per PCR. This equates to a final concentration of 0.2 µM.
Due to the small volumes of primers used per rxn, we suggest making batch PCR mixes (e.g. 5, 10, 20, 32 rxns) and aliquoting 20 µL of the complete PCR mix into PCR tubes before adding DNA template to each PCR as shown in the table below.
An additional 10% of all components has been added to the PCR mix table below to compensate for any pipetting errors during aliquoting.
If you wish to alter the volume of primer used, or to use any PCR additives, add or subtract the additional volume from the volume of PCR-grade water.
Components
1 rxn (µL)
5 rxn* (µL)
10 rxn* (µL)
20* (µL)
32* (µL)
PCR grade water
13.2
56
145
290.4
464.64
5x Mastermix
4
22
44
88
140.8
Forward primer
0.4
2.2
4.4
8.8
14.1
Reverse primer
0.4
2.2
4.4
8.8
14.1
DNA Template (total)
2
10
20
40
70.4
Final volume
20
100
200
400
640
Excess
0
10
20
40
70.4
*Volumes contain 10% extra reagents to compensate for any potential pipetting errors.
Primer sequences
Forward: ITS1F: 5’-CTTGGTCATTTAGAGGAAGTAA-3’
Reverse: ITS4: 5’-TCCTCCGCTTATTGATATGC-3’
Expected amplicon length
500-1000 bp, depending on species.
Components
Each primer pair contains two tubes:
Forward primer (10 µM, 200 µL, in low TE buffer)
Reverse primer (10 µM, 200 µL, in low TE buffer)
Storage & Stability
Suitable for use without refrigeration or ice. The primers should be stable at room temperature for up to six months if unopened or if kept completely sterile, but to extend stability store at 4 ºC or -20 ºC. Once opened store at 4 ºC short-term, or at -20 ºC for long-term storage. Do not store in direct sunlight.
