Two-step genomic sequence comparison strategy to design Trichoderma strain-specific primers for quantitative PCR.

Survival of inoculated fungal strains in a brand new surroundings performs a important position in practical efficiency, however few research have centered on strain-specific quantitative PCR (qPCR) strategies for monitoring useful fungi.

On this research, the Trichoderma guizhouense pressure NJAU 4742 (remodeled with the gfp gene and named gfp-NJAU 4742), which reveals a growth-promoting impact via phytohormone manufacturing and pathogen antagonism, was chosen as a mannequin to design strain-specific primer pairs utilizing two steps of genomic sequence comparability to detect its abundance in soil.

After a second comparability with the carefully associated species T. harzianum CBS 226-95 to additional differentiate the strain-specific fragments that had proven no homology to any sequence deposited within the databases used within the first comparability, ten primer pairs had been designed from the entire genome.

In the meantime, three primer pairs, P11, P12 and P13, had been additionally designed from the inserted fragment containing the gfp gene. After verification testing with three varieties of subject soils, primer pairs P6, P7 and P8 had been additional chosen by comparability with P11, P12 and P13.

A sensible check utilizing a pot experiment confirmed that steady colonization of gfp-NJAU 4742 in pepper rhizosphere soil could possibly be detected utilizing primer pairs P6 and P7, displaying no important distinction from the outcomes of primers P11 and P12.

Therefore, the technique described right here for designing fungal-strain-specific primers might theoretically be used for some other fungi for which the entire genome sequence is on the market in a database, and the qPCR methodology developed will also be used to additional monitor the inhabitants dynamics of various strains primarily based on the designed primers.

Quantitative PCR primer design impacts quantification of dsRNA-mediated gene knockdown.

RNA interference (RNAi) is a robust software for learning features of candidate genes in each mannequin and nonmodel organisms and a promising approach for therapeutic functions. Profitable software of this system depends on the accuracy and reliability of strategies used to quantify gene knockdown.

With the limitation within the availability of antibodies for detecting proteins, quantitative PCR (qPCR) stays the popular technique for quantifying goal gene knockdown after dsRNA therapy. We evaluated how qPCR primer binding web site and goal gene expression ranges have an effect on quantification of intact mRNA transcripts following dsRNA-mediated RNAi.

Two-step genomic sequence comparison strategy to design Trichoderma strain-specific primers for quantitative PCR.

The usage of primer pairs focusing on the mRNA sequence inside the dsRNA goal area did not reveal a major lower in goal mRNA transcripts for genes with low expression ranges, however not for a extremely expressed gene. Against this, important knockdown was detected in all instances with primer pairs focusing on the mRNA sequence extending past the dsRNA goal area, whatever the expression ranges of the goal gene.

Our outcomes counsel that at the least for genes with low expression ranges, quantifying the effectivity of dsRNA-mediated RNAi with primers amplifying sequences fully contained within the dsRNA goal area ought to be prevented because of the danger of false-negative outcomes. As an alternative, primer pairs extending past the dsRNA goal area of the mRNA transcript sequences ought to be used for correct and dependable quantification of silencing effectivity.

Combining bioinformatics and standard PCR optimization technique for one-time design of high-specificity primers for WRKY gene household utilizing unigene database.

Gene households, just like the conserved transcription issue households, evolve via gene duplications and share reasonable similarity between member genes. Lack of genomic information makes it tough to design high-specificity primers to the goal genes.

Moreover, many primers under-perform in extremely delicate assays like quantitative PCR as a result of problems with thermodynamic nature, thereby growing the fee and time for evaluation. A strategy involving intra-species and inter-generic bioinformatic sequence comparability mixed with thermodynamic estimation of primer efficiency was used for one-time design of gene particular primers for various WRKYs, Mitogen Activated Protein-kinases and N-methyltransferases of Coffea canephora with out the help of genome sequence assets.

Out of a complete 37 primer units together with 31 pairs of primers for WRKY from 34 mined WRKY Unigenes/ESTs and 6 pairs for genes coding for MAP kinases and NBS-LRR proteins, 32 units exhibited excessive specificity of amplification upon genome evaluation in addition to within the high-resolution soften evaluation.

Moreover, PCR optimization strategies-both in silico and experimental-indicated a superior efficiency of the primer units for various functions like quantitative PCR and speedy amplification of cDNA ends. Just one set of primer resulted in mis-priming upon affirmation by DNA sequencing of the cloned amplicons.

The intra-species variations and inter-generic similarities guarantee excessive specificity of primers in all instances studied. The process allowed design of primers for the use in numerous downstream functions with excessive efficiency, specificity, yield and ease-of-use.

Primer design and amplification efficiencies are essential for reliability of quantitative PCR research of caffeine biosynthetic N-methyltransferases in espresso.

Primers having suboptimal amplification efficiencies had been proven to falsely characterize fold change expression of the N-methyltransferases gene household concerned in caffeine biosynthesis in Coffea canephora.

To check this phenomenon, the position of stability of the interior reference gene, in addition to the amplification effectivity correction of the primers was investigated. GAPDH and Ubiquitin exhibited a great stability for learning the ontogeny of endosperm tissue, in addition to the leaf transcriptome throughout stress from salicylic acid, methyl jasmonate, PEG-mediated drought and sudden publicity to mild.

Ubiquitin manifested low variation in Cq beneath all these stress regimes and in endosperm ontogeny with 30.1-30.9 in one of the best dataset and 28.8-30.9 in probably the most deviating dataset. It was noticed that issues arising as a result of improper amplification effectivity of the goal or reference genes or each may result in misinterpretation of gene expression ranges.

Quantitative RT-PCR carried out at a sub-optimal effectivity of GAPDH reference gene at 1.68 led to the defective interpretation of two.007 folds upregulation by the 2-ΔΔCt technique and 1.705 folds upregulation by Effectivity technique for the primary NMT (Xanthosine methyltransferase), which truly is repressed throughout darkish acclimatization of espresso vegetation.

Effectivity correction improved the reliability of the expression information and likewise indicated a downregulation of this gene by 0.485 folds and 0.474 folds utilizing 2-ΔΔCt and E technique, respectively, in concordance to earlier studies.

The egg drop syndrome (76) PCR kit

PCR-V044-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of The egg drop syndrome (76)

The egg drop syndrome (76) PCR kit

PCR-V044-96D 100T
EUR 521.5
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of The egg drop syndrome (76)

PCR Mix

L5051100 2.5 ml
EUR 107
Description: Premade ready to use kits will always come in handy. Get your experiment done right form the first try by using a validated kit with perfectly balanced reagents proportions and compatibility and by following a clear protocol.

2×Taq PCR Master Mix(with dye)

K1034-1 1 ml
EUR 73

Ready? PCR Mix

M1127-1000
EUR 512

Ready? PCR Mix

M1127-200
EUR 191

Rigor? PCR Mix

M1132-200
EUR 321

Breeze? PCR Mix

M1134-200
EUR 267

Distant? PCR Mix

M1136-200
EUR 294

Advance? PCR Mix

M1139-200
EUR 376

Egg Drop Syndrome RT PCR kit

RTq-V026-100D 100T
EUR 628.5
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Description: A Real-Time PCR kit for detection of Egg Drop Syndrome .

Egg Drop Syndrome RT PCR kit

RTq-V026-150D 150T
EUR 701
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Description: A Real-Time PCR kit for detection of Egg Drop Syndrome .

Egg Drop Syndrome RT PCR kit

RTq-V026-50D 50T
EUR 532.8
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Description: A Real-Time PCR kit for detection of Egg Drop Syndrome .

dNTP Mix (1 ml)

9002-1
EUR 153

Individual Reaction Mix 1

G065-1 200 reactions
EUR 167

PCR-EZ D-PCR MASTER MIX

BS294 100RXN, 100prep
EUR 93.5
  • Product category: PCR Related/PCR Premix (dPCR)/dPCR Premix + Taq

amfiSure PCR Master Mix

P0311-010 2x50 rxns
EUR 126

amfiSure PCR Master Mix

P0311-025 5X50 rxns
EUR 165

amfiSure PCR Master Mix

P0311-050 10X50 rxns
EUR 219

amfiSure PCR Master Mix

P0311-100 10x1ml
EUR 286

amfiSure PCR Master Mix

P0311-125 15X50 rxns
EUR 278

amfiSure PCR Master Mix

P0311-200 20x1ml
EUR 469

amfiSure PCR Master Mix

P0311-250 50x50 rxns
EUR 752

amfiSure PCR Master Mix

P0311-500 100x50 rxns
EUR 1258

amfiXpand PCR Master Mix

P0331-010 2X50 rxns
EUR 151

amfiXpand PCR Master Mix

P0331-025 5X50 rxns
EUR 273

amfiXpand PCR Master Mix

P0331-050 10X50 rxns
EUR 448

amfiXpand PCR Master Mix

P0331-250 50x50 rxns
EUR 1873

Ready? PCR Mix-Dye

M1128-1000
EUR 512

Ready? PCR Mix-Dye

M1128-200
EUR 191

Image Ready? PCR Mix

M1129-200
EUR 294

Robust Ready? PCR Mix

M1130-1000
EUR 615

Robust Ready? PCR Mix

M1130-200
EUR 213

Rigor? PCR Mix-Dye

M1133-200
EUR 321

Breeze? PCR Mix-Dye

M1135-200
EUR 267

Distant? PCR Mix-Dye

M1137-200
EUR 294

Image Distant? PCR Mix

M1138-200
EUR 321

Advance? PCR Mix-Dye

M1140-200
EUR 376

Fire Start? PCR Mix

M1141-200
EUR 376

Whole Blood PCR Mix

M1143-200
EUR 321

Egg Drop Syndrome One-Step PCR kit

Oneq-V026-100D 100T
EUR 747.4
  • Contact us in order to know the reactivity of the kit.
Description: Real Time PCR Kit is a screening assay for a rapid and accurate detection of Egg Drop Syndrome.

Egg Drop Syndrome One-Step PCR kit

Oneq-V026-150D 150T
EUR 838.75
  • Contact us in order to know the reactivity of the kit.
Description: Real Time PCR Kit is a screening assay for a rapid and accurate detection of Egg Drop Syndrome.

Egg Drop Syndrome One-Step PCR kit

Oneq-V026-50D 50T
EUR 628.5
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Description: Real Time PCR Kit is a screening assay for a rapid and accurate detection of Egg Drop Syndrome.

2x GoldStar Best PCR Master Mix (with Dyes), 20 ul/rxn

W0655-1 NULL
EUR 0

MyTaq Blood PCR Mix, 2x

BIO-25053/S Sample Ask for price

MyTaq Blood PCR Mix, 2x

BIO-25054 250 Reactions Ask for price

amfiSure Prime PCR Master Mix

P1311-025 5X50 rxns
EUR 161

amfiSure Prime PCR Master Mix

P1311-050 10X50 rxns
EUR 219

amfiSure Prime PCR Master Mix

P1311-125 15X50 rxns
EUR 273

amfiSure Prime PCR Master Mix

P1311-250 50x50 rxns
EUR 739

amfiSure Prime PCR Master Mix

P1311-500 100x50 rxns
EUR 1238

amfiSure Advanced PCR Master Mix

P2311-025 5X50 rxns
EUR 165

amfiSure Advanced PCR Master Mix

P2311-050 10X50 rxns
EUR 223

amfiSure Advanced PCR Master Mix

P2311-125 15X50 rxns
EUR 282

amfiSure Advanced PCR Master Mix

P2311-250 50x50 rxns
EUR 752

amfiSure Advanced PCR Master Mix

P2311-500 100x50 rxns
EUR 1279

Robust Ready? PCR Mix-Dye

M1131-1000
EUR 615

Robust Ready? PCR Mix-Dye

M1131-200
EUR 213

Fire Start? PCR Mix-Dye

M1142-200
EUR 376

mRNAExpress Tail PCR mix (5uM)

MR-TAIL-PR 100 ul
EUR 164
  • Category: MicroRNA Tools

The egg drop syndrome (76) RT PCR kit

RTq-V044-100D 100T
EUR 628.5
  • Contact us in order to know the reactivity of the kit.
Description: A Real-Time PCR kit for detection of The egg drop syndrome (76) .

The egg drop syndrome (76) RT PCR kit

RTq-V044-150D 150T
EUR 701
  • Contact us in order to know the reactivity of the kit.
Description: A Real-Time PCR kit for detection of The egg drop syndrome (76) .

The egg drop syndrome (76) RT PCR kit

RTq-V044-50D 50T
EUR 532.8
  • Contact us in order to know the reactivity of the kit.
Description: A Real-Time PCR kit for detection of The egg drop syndrome (76) .

PRICE DROP

110-012 4 x 1000 µl
EUR 133

PRICE DROP

110-012L 5x 4 x 1000 µl
EUR 518

PRICE DROP

110-012XL 10x 4 x 1000 µl
EUR 939

The egg drop syndrome (76) One-Step PCR kit

Oneq-V044-100D 100T
EUR 747.4
  • Contact us in order to know the reactivity of the kit.
Description: Real Time PCR Kit is a screening assay for a rapid and accurate detection of The egg drop syndrome (76).

The egg drop syndrome (76) One-Step PCR kit

Oneq-V044-150D 150T
EUR 838.75
  • Contact us in order to know the reactivity of the kit.
Description: Real Time PCR Kit is a screening assay for a rapid and accurate detection of The egg drop syndrome (76).

The egg drop syndrome (76) One-Step PCR kit

Oneq-V044-50D 50T
EUR 628.5
  • Contact us in order to know the reactivity of the kit.
Description: Real Time PCR Kit is a screening assay for a rapid and accurate detection of The egg drop syndrome (76).

Taq 5x PCR Mix 12.5 mM

BT10402 250rxn
EUR 109
Description: High quality Taq polymerase for different PCR variations and downstream applications.

Taq 2x PCR Mix 100 rxn

BA01501 100rxn
EUR 94
Description: High quality Taq polymerase for different PCR variations and downstream applications.

HotTaq 2x PCR Mix 100 rxn

BA01503 100rxn
EUR 120
Description: High quality HotTaq polymerase for different PCR variations and downstream applications.

RedTaq 2x PCR Mix 100 rxn

BA01507 100rxn
EUR 146
Description: High quality RedTaq polymerase for different PCR variations and downstream applications.

amfiSure ONE PCR Master Mix(2X)

P7000-005 5x1 ml
EUR 219

amfiSure ONE PCR Master Mix(2X)

P7000-010 10x1 ml
EUR 317

amfiSure ONE PCR Master Mix(2X)

P7000-050 50x1 ml
EUR 1211

amfiSure ONE PCR Master Mix(2X)

P7000-100 100x1 ml
EUR 2102

AXYPREP MAG PCR NORMALIZER KIT- 1 ML - (SAMPLE SIZE)

MAG-PCR-NM-1 1/pk
EUR 73
Description: Bioscience Mag Beads; Magnetic PCR Normaliser

AXYPREP MAG PCR CLEAN UP KIT- 1 ML - (SAMPLE SIZE)

MAG-PCR-CL-1 1/pk
EUR 58
Description: Bioscience Mag Beads; Magnetic PCR Clean Up

Poliovirus 1 (PV1) PCR kit

PCR-H659-48R 50T
EUR 524.4
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Description: An conventional PCR kit for detection of Poliovirus 1 (PV1)

Poliovirus 1 (PV1) PCR kit

PCR-H659-96R 100T
EUR 669.4
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Description: An conventional PCR kit for detection of Poliovirus 1 (PV1)

Psittacid herpesvirus 1 PCR kit

PCR-V014-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of Psittacid herpesvirus 1

Psittacid herpesvirus 1 PCR kit

PCR-V014-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of Psittacid herpesvirus 1

Equine herpesvirus 1 PCR kit

PCR-V144-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of Equine herpesvirus 1

Equine herpesvirus 1 PCR kit

PCR-V144-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of Equine herpesvirus 1

Bovine herpesvirus 1 PCR kit

PCR-V182-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of Bovine herpesvirus 1

Bovine herpesvirus 1 PCR kit

PCR-V182-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of Bovine herpesvirus 1

Bluetongue virus 1 PCR kit

PCR-V226-48R 50T
EUR 524.4
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Description: An conventional PCR kit for detection of Bluetongue virus 1

Bluetongue virus 1 PCR kit

PCR-V226-96R 100T
EUR 669.4
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Description: An conventional PCR kit for detection of Bluetongue virus 1

Smart 2X PCR pre-mix Pfu (30ul)

9K-002-0038 30uL, 30uL
EUR 581.14
  • Product category: PCR Related/PCR Premix/PCR Premix + Pfu

HotStart PCR Master mix (2X, Green Dye)

BS9291 5X1mL, 5ml
EUR 209.5
  • Product category: PCR Related/PCR Premix (qPCR)/qPCR Premix + Taq

HotStart PCR Master mix (2X, Green Dye)

BS9292 1ml
EUR 96.4
  • Product category: PCR Related/PCR Premix (qPCR)/qPCR Premix + Taq

Taq PCR Master Mix (2X, Blue Dye)

BS9295 1ml
EUR 84.8
  • Product category: PCR Related/PCR Premix (qPCR)/qPCR Premix + Taq

Taq PCR Master Mix (2X, Blue Dye)

BS9296 5ml
EUR 189.2
  • Product category: PCR Related/PCR Premix (qPCR)/qPCR Premix + Taq

Taq PCR Master Mix (2X, Red Dye)

BS9297 1ml
EUR 84.8
  • Product category: PCR Related/PCR Premix (qPCR)/qPCR Premix + Taq

Taq PCR Master Mix (2X, Red Dye)

BS9298 5ml
EUR 189.2
  • Product category: PCR Related/PCR Premix (qPCR)/qPCR Premix + Taq

2×Taq PCR Master Mix(with dye)

K1034-100 100×1 ml
EUR 792

2×Taq PCR Master Mix(with dye)

K1034-20 20×1 ml
EUR 282

2×Taq PCR Master Mix(with dye)

K1034-5 5×1 ml
EUR 119

2×Taq PCR Master Mix(with dye)

K1034-50 50×1 ml
EUR 514

amfiFusion High Fidelity PCR Master Mix(2X)

P0342-010 2x50 rxns
EUR 253

amfiFusion High Fidelity PCR Master Mix(2X)

P0342-025 5X50 rxns
EUR 455

amfiFusion High Fidelity PCR Master Mix(2X)

P0342-050 10X50 rxns
EUR 779

amfiFusion High Fidelity PCR Master Mix(2X)

P0342-100 20x50 rxns
EUR 1333

amfiSure Ultra Fidelity PCR Master Mix(2X)

P0346-001 1ml
EUR 280

amfiSure Ultra Fidelity PCR Master Mix(2X)

P0346-002 2x1ml
EUR 482

amfiSure Ultra Fidelity PCR Master Mix(2X)

P0346-005 5x1ml
EUR 828

amfiSure Ultra Fidelity PCR Master Mix(2X)

P0346-010 10x1ml
EUR 1360

Reverse Transcriptase RT PCR Master Mix (5X)

MB1002-100reactions 100 reactions
EUR 514

Reverse Transcriptase RT PCR Master Mix (5X)

MB1002-200reactions 200 reactions
EUR 746

Reverse Transcriptase RT PCR Master Mix (5X)

MB1002-25reactions 25 reactions
EUR 189

MULTIPLEX KIT PCR MASTITIS PCR kit

PCR-MPX218-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of MULTIPLEX KIT PCR MASTITIS

MULTIPLEX KIT PCR MASTITIS PCR kit

PCR-MPX218-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of MULTIPLEX KIT PCR MASTITIS

Herpes simplex type 1 PCR kit

PCR-H426-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of Herpes simplex type 1

Herpes simplex type 1 PCR kit

PCR-H426-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of Herpes simplex type 1

Dengue virus type 1 PCR kit

PCR-H521-48R 50T
EUR 524.4
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Description: An conventional PCR kit for detection of Dengue virus type 1

Dengue virus type 1 PCR kit

PCR-H521-96R 100T
EUR 669.4
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Description: An conventional PCR kit for detection of Dengue virus type 1

Canine adenovirus type 1 PCR kit

PCR-V077-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of Canine adenovirus type 1

Canine adenovirus type 1 PCR kit

PCR-V077-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of Canine adenovirus type 1

Caprine herpes virus 1 PCR kit

PCR-V312-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of Caprine herpes virus 1

Caprine herpes virus 1 PCR kit

PCR-V312-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of Caprine herpes virus 1

Mouse parvovirus type 1 PCR kit

PCR-V324-48D 50T
EUR 425.8
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Description: An conventional PCR kit for detection of Mouse parvovirus type 1

Mouse parvovirus type 1 PCR kit

PCR-V324-96D 100T
EUR 521.5
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Description: An conventional PCR kit for detection of Mouse parvovirus type 1

Borna disease viruses 1 PCR kit

PCR-V605-48R 50T
EUR 524.4
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Description: An conventional PCR kit for detection of Borna disease viruses 1

Borna disease viruses 1 PCR kit

PCR-V605-96R 100T
EUR 669.4
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of Borna disease viruses 1

Fast Probe Master Mix (500 rxn)

31005-1 5x1ML
EUR 466
Description: Minimum order quantity: 1 unit of 5x1ML

2xEs Taq Master Mix (with Dyes)

W0690-1 NULL
EUR 0

MULTIPLEX KIT PCR Babesia & Theileria PCR kit

PCR-MPX401-48D 50T
EUR 425.8
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of MULTIPLEX KIT PCR Babesia & Theileria

MULTIPLEX KIT PCR Babesia & Theileria PCR kit

PCR-MPX401-96D 100T
EUR 521.5
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of MULTIPLEX KIT PCR Babesia & Theileria

Primer Mix for Junction PCR to Confirm AAVS1 Integration Site (compatible with GE620A-1, GE622A-1, GE624A-1 and CAS620A-1), 50ul at 5uM for each primer set

GE640PR-1 50 reactions
EUR 159
  • Category: Gene Editing

Drop Delay Calibration Particles

DDCP-70-2 2 mL
EUR 283
Description: Please reffer to the technical data sheet for more detail information for this item. Our dedicated team would be happy to assist you via live chat, email or phone.

Drop Delay Calibration Particles

DDCP-70-20 20 mL
EUR 1918
Description: Please reffer to the technical data sheet for more detail information for this item. Our dedicated team would be happy to assist you via live chat, email or phone.

Human T-lymphotropic Virus 1 PCR kit

PCR-H657-48R 50T
EUR 524.4
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of Human T-lymphotropic Virus 1

Human T-lymphotropic Virus 1 PCR kit

PCR-H657-96R 100T
EUR 669.4
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of Human T-lymphotropic Virus 1

Equine rotavirus strain H-1 PCR kit

PCR-V161-48R 50T
EUR 524.4
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of Equine rotavirus strain H-1

Equine rotavirus strain H-1 PCR kit

PCR-V161-96R 100T
EUR 669.4
  • Contact us in order to know the reactivity of the kit.
Description: An conventional PCR kit for detection of Equine rotavirus strain H-1

Therefore, effectivity correction of the primers having suboptimal efficiencies is an absolute prerequisite for the correct calculation of fold change utilizing quantitative RT-PCR.

 

 

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