Overview

• Detection kits for Vibrio cholerae
• Lyophilized format designed for room temperature shipping
• Available in TaqMan format for analysis

Norgen’s Vibrio cholerae TaqMan PCR Lyophilized Kit is designed for the detection of Vibrio cholerae specific DNA in a real-time PCR based on the use of TaqMan® technology. The lyophilized format is designed to ship the kit at ambient temperature.

Norgen’s Vibrio cholerae TaqMan Lyophilized Probe/Primer and Control Set is designed for the detection of Vibrio cholerae specific DNA in a real-time PCR based on the use of TaqMan® technology. The lyophilized format is designed to ship the kit at ambient temperature.

Details

Supporting Data

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Storage Conditions and Product Stability

• All kit components should be stored at -20°C upon arrival.
• Once reconstituted, repeated thawing and freezing (>2 times) of the Master Mix and Positive Control should be avoided, as this may affect the performance of the assay. If the reagents are to be used only intermittently, they should be frozen in aliquots.
• All kit components can be stored for 2 years after the date of production without showing any reduction in performance.

Component	Cat. TM38550L (100 preps)	Cat. TM38510L (100 preps)
MDx TaqMan 2X PCR Master Mix (Lyo)	4 x 350 μL	–
Vibrio cholerae Primer & Probe Mix (Lyo)	280 μL	280 μL
Vibrio cholerae Positive Control (Lyo)	150 μL	150 μL
Nuclease-Free Water (Negative Control)	3 x 1.25 mL	1 x 1.25 mL
Product Insert	1	1

Endonucleases Non-Specific, HL-SAN

OverView

HL-SAN efficiently removes nucleic acids from buffers typically used in protein purification. Due to its high salt tolerance, it is the obvious choice for host-cell DNA removal in settings where salt is added to reduce aggregation. Especially efficient for removing nucleic acids from proteins with high affinity for DNA and RNA. Proven performance during lysis and early stages of protein purification processes, as well as high-salt eluates. Cold-adapted enzyme with excellent performance also at ambient temperatures and during over-night digestion at 4°C.

Recommended applications:

Complete removal of DNA 

Viscosity reduction

Key advantages with this Salt Active Nuclease:

• Non specific endonuclease
• Optimum activity at high salt concentration (0.5 M NaCl)
• Active at low temperatures (20% at 6ºC)
• Easily inactivated
• Broad pH range
• Temperature stable

Figures

Figure 1. Optimum activity in solutions with high salinity

HL-SAN has optimum activity at ∼0.5 M NaCl, but operates at a broad range of [NaCl] and [KCl]. The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 5 mM MgCl₂ with varying [NaCl] or [KCl]. The maximum activity was set to 100%.

Figure 2. Temperature and activity

HL-SAN has optimum activity at ~35°C, but works over a broad temperature range (20% activity at 10°C and 50°C). The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5 containing 5 mM MgCl₂ and 0.5 M NaCl.

Fig 3. The effect of MgCl₂ and MnCl₂ concentration on the HL-SAN activity.

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 0.5 M NaCl and with varying concentrations of MgCl₂ or MnCl₂. The activity of the sample containing 5 mM MgCl₂ was set to 100%.

Figure 4. HL-SAN activity vs pH/[NaCl]

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer with different pHs and different concentrations of NaCl. All buffers contained 5 mM MgCl₂. The nature of the buffer was pH-dependent, but generally the NaCl-optimum was the same in all buffers/pHs. The exception was etanolaminbuffer at pH 9 and pH 9.5 in which the NaCl-optimum was shifted to the left (not shown).

Figure 5. Buffer composition affects substrate preference

Without NaCl, the specificity towards ssDNA and dsDNA is similar. At 0.5 M NaCl, the activity towards dsDNA increases, while the activity towards ssDNA is unaffected.

Figure 6. HL-SAN digests ssDNA to ~5-13 nt, and dsDNA to ~5-7 nt

The size of the end products from ssDNA varies from ~5-13 nt, while dsDNA is digested to around ~5-7 nt. The size of the end products seems to depend on the DNA sequence. Substrates 1 and 2 were ssDNA with different sequences and substrates 3 and 4 were dsDNA with similar sequences but with a FAM-label at different ends. Substrate 5 was dsDNA with the same sequence as substrate 3 and 4 but with a FAM-label at both ends.

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Figure 7. HL-SAN activity decreases with increasing concentrations of glycerol

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 5 mM MgCl₂, 0.5 M NaCl and with increasing concentrations of glycerol. The activity of the control not containing glycerol was set to 100%.

Figure 8. The activity of HL-SAN at different concentrations of imidazole

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 5 mM MgCl₂, 0.5 M NaCl and with varying concentrations of imidazole. The activity of the control not containing imidazole was set to 100%.

HL-SAN efficiently removes nucleic acids from buffers typically used in protein purification. Due to its high salt tolerance, it is the obvious choice for host-cell DNA removal in settings where salt is added to reduce aggregation. Especially efficient for removing nucleic acids from proteins with high affinity for DNA and RNA. Proven performance during lysis and early stages of protein purification processes, as well as high-salt eluates. Cold-adapted enzyme with excellent performance also at ambient temperatures and during over-night digestion at 4°C.

Introduction

This product is suitable for rapid extraction of total DNA from tissue, cells, blood, saliva, swabs, blood spots, semen and other clinical samples. DNA can be used directly for PCR, quantitative PCR, Southern Blot, test of virus DNA and so on.

Details

Specifications

Features	Specifications
Main Functions	Isolation total DNA from tissue / blood / body fluid / swab /dry blood spots
Applications	PCR, qPCR, southern bolt and virus detection, etc.
Purification method	Mini spin column
Purification technology	Silica technology
Process method	Manual (centrifugation or vacuum)
Sample type	Tissue, cell, blood, saliva, swab, blood spot, semen and other clinical samples
Sample amount	Solid tissue : 1-10mg, Anticoagulant blood : 200µl
Yield	1 – 15µg
Elution volume	≥20μl
Time per run	30 – 60 minutes
Liquid carrying volume per column	750µl
Binding yield of column	100µg

Principle

This product is based on silica Column purification. The sample is lysed and digested with lysate and protease, DNA is released into the lysate. Transfer to an adsorption column. Nucleic acid is adsorbed on the membrane, while protein is not adsorbed and is removed with filtration. After washing proteins and other impurities, nucleic acid was finally eluted with low-salt buffer (10mm Tris, pH9.0, 0.5mm EDTA).

Advantages

• High quality DNA – meet a variety of downstream applications, including PCR, qPCR, enzyme digestion, hybridization, etc.
• Fast – without separation of leukocytes, organic extraction or ethanol precipitation
• Simple – all nucleic acids can be obtained by direct digestion
• Wide applicability – It can handle various liquid samples, animal tissues and cultured cells

Kit Contents

Contents	IVD3018
Purification Times	100
HiPure DNA Mini Columns I	100
2ml Collection Tubes	2 x 100
Buffer ATL	60 ml
Buffer AL	60 ml
Buffer GW1	44 ml
Buffer GW2	50 ml
Proteinase K	60 mg
Protease Dissolve Buffer	5 ml
Buffer AE	15 ml

Storage and Stability

Proteinase K should be stored at 2–8°C upon arrival. However, short-term storage (up to 12 weeks) at room temperature (15–25°C) does not affect their performance. The remaining kit components can be stored at room temperature (15–25°C) and are stable for at least 18 months under these conditions. The entire kit can be stored at 2–8°C, but in this case buffers should be redissolved before use. Make sure that all buffers are at room temperature when used.

Experiment Data

This product is suitable for rapid extraction of total DNA from tissue, cells, blood, saliva, swabs, blood spots, semen and other clinical samples. DNA can be used directly for PCR, quantitative PCR, Southern Blot, test of virus DNA and so on.