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Custom DNA Sequencing(Inquire for pricing)
 
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Code: Custom DNA Sequencing
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CUSTOM DNA SEQUENCING DESCRIPTION

 

CUSTOM DNA SEQUENCING DESCRIPTION

For more information, please click on the following:

Pricing, Online form, Universal Primers ,

Submission Template Requirement , Turnaround time and Custom primers (Oligo synthesis) for Sequencing




PRIMER EXTENSION SEQUENCING SERVICE

Result:

Up to 1100 bp sequence data per reaction.

Starting Material:

Provided templates from customer.

Provided primers from customer

Custom primers can be synthesized at our company for faster results.

The following universal primers are available to you at no cost: Please click here Universal Primers

Method:

Primer-extension by using big-dye chemistry or other related chemistry from PE.

If the sequence data is unsatisfactory, the sequencing reaction is repeated with different condition. The better result will be sent to the customer.

 

DIRECT PCR PRODUCT SEQUENCING SERVICE

 

Result:

Up to 1100 bp sequence data per reaction.

Starting Material:

Purified PCR products from customer.

Provided primers from customer.

Method:

Primer-extension by using big-dye chemistry or other related chemistry form PE.

If the sequence data is unsatisfactory, the sequencing reaction is repeated with different condition. The better result will be sent to the customer.

 

SINGLE STRANDED DNA SEQUENCING SERVICE

 

Result:

Consensus sequence with > 99% accuracy.

A complete report of sequencing strategies and data.

Starting Material:

Provided templates from customer.

Customer-provided initial primers.

Custom primers can be synthesized at our company for faster results.

The following universal primers are available to you at no cost: Please click here Universal Primers

Method:

Design and synthesize internal primers at the optimal interval for better overlapping.

Single stranded coverage of the template through multiple sequencing reactions.

Resolve most problematic regions.

The consensus sequence is generated with accuracy greater than 99 %.


 
DOUBLE STRANDED DNA SEQUENCING SERVICE


Result:

Finished consensus sequence with greater than 99.99% of accuracy.

A complete report of sequencing strategies and data.

Starting Material:

Templates from customer.

Customer-provided primers.

Custom primers can be synthesized at our company for faster results.

The following universal primers are available to you at no cost: Please click here Universal Primers

Method:

Design and synthesize internal primers.

Double stranded coverage of the template through multiple sequencing reactions.

Assemble and edit the contigs.

The problematic regions are completely determined.

The consensus sequence is generated with accuracy greater than 99.99%.

 

BAC, PAC AND P1 ENDS SEQUENCING SERVICE

 

Result

400-600 bp sequence data

Starting Material:

Provided templates form customer.

Provided primers from customer

Method:

Proprietary protocols in conjunction with big-dye chemistry or other related chemistry form PE.

If the sequence data is unsatisfactory, the sequencing reaction is repeated with different condition. The better result will be sent to the customer.

 

cDNA/EST LIBRARY SEQUENCING SERVICE

 

Result:

up to 1100bp sequence data per reaction

Starting Material:

Provided bacterial cultures or templates from customer.

Customer-provided primers.

Custom primers can be synthesized at our company for faster results.

The following universal primers are available to you at no cost: Please click here Universal Primers .

If the sequence data is unsatisfactory, the sequencing reaction is repeated with different condition. The better result will be sent to the customer.

Method:

Primer-extension by using big-dye kit or other related chemistry form PE.

Use automated liquid sample handling instrument to prepare sequencing reactions.

If the sequence data is unsatisfactory, the sequencing reaction is repeated with different condition. The better result will be sent to the customer.

The samples are cataloged and stored at -20 C.

 

 

FDA SUBMISSION SEQUENCING SERVICE

 

Result:

Finished consensus sequence with 100 % of accuracy.

A complete report of sequencing strategies and data.

Starting Material:

Templates from customer.

Customer-provided primers.

Custom primers can be synthesized at our company for faster results.

The following universal primers are available to you at no cost: Please click here Universal Primers. .

Method:

Use big-dye chemistry or other related chemistry from PE.

Complete double stranded coverage of the template through 4 fold-redundancy sequencing.

Resolve all problematic regions.

Assemble and edit the final sequence.

 

PLASMID DNA PREPARATION SERVICE

 

Result:

Purified plasmids suitable for automated sequencing, transfection, and other molecular methodologies.

Starting Material:

Provided bacterial cultures from customer.

Method:

Proprietary protocols or commercial kits.

 

PCR PRODUCT PREPARATION SERVICE

 

Result:

Purified PCR fragments.

Starting Material:

Crude PCR products.

Expected size of the PCR product to be isolated and purified.

Method:

Proprietary protocols or commercial kits.

 

Submission Template Requirement

 

Plasmid DNA Preparation

The quality of template DNA is extremely important to obtain good sequence data. The following methodologies are recommended for preparing plasmid DNA for automated sequencing: Cessium chloride (CsCl) banding, Qiagen plasmid kit, Wizard plus plasmid kit, and other commercial kits available. Magnesium ions are essential for DNA polymerase activity. Template DNA and primer should be resuspended in water or a buffer containing no more than 0.1 mM EDTA. Introduction of large amounts of EDTA in template DNA or primer will result in weak signals or short reads. The optimal concentration of the template should be at 0.2 ug/ul.

PCR Product Preparation

The purity of the PCR product is very crucial for obtaining good sequence data. Any PCR primers and/or dNTPs carryover in the PCR product will adversely affect the quality of the sequence data. If the PCR product has a unique band, it can be purified by using size exclusion method, like PCR purification kit from Qiagen and Pharmacia. If the PCR product has more than one bands, the PCR product should be ran on the agarose gel and isolate out the band that is needed to do sequencing. The band can be purified by using the kit from Centricon or Qiagen. The purified PCR products can be quantitated either on the gel or spectrophotometer.

QC Your Templates

It is very important to measure the concentration of your template accurately, either on the agarose gel or by spectrophotometer. The #1 factor that causes poor sequencing results is due to inaccurate concentration. So, please verify the concentration of your template accurately.


Recommended DNA Template and Primer Quantities

 

 Product

Amount

Concentration

 

PCR Product DNA

 

 

 

100-500 bp

150 ng

20 ng/ul

 

500-1000bp

200 ng

20 ng/ul

 

1000-2000bp

300 ng

20 ng/ul

 

Single stranded

600 ng

100 ng/ul

 

Double stranded

1.5 ug

200 ng/ul

 

Cosmid, BAC, P1

4 ug

200 ng/ul

 

Custom primer

100 picomoles

10 picomoles/ul

 

 

Please see below for more detail information:

Recommended DNA Template and Primer Quantities for Each Reaction
DNA and Primer Separated

DNA

Concentration

Amount

2x Amount

PCR product DNA

5-20ng/l

50ng

100ng

Single stranded plasmid

50-100ng/l

250ng

500ng

Double stranded plasmid

50-200ng/l

500ng

1000ng

BAC

100-200ng/l

2000ng

4000ng

Primer

10pmol/l

5l

10l

We recommend 2x amount of DNA in case we need to repeat the reactions.

DNA and Primer Combined

DNA

DNA

Primer

Total Vol.

2X DNA

2X Primer

2X Total Vol.

PCR product DNA

50ng

10pmoles

10uL

100ng

20pmoles

20uL

Single stranded plasmid

250ng

10pmoles

10uL

500ng

20pmoles

20uL

Double stranded plasmid

500ng

10pmoles

10uL

1000ng

20pmoles

20uL

BAC

2000ng

10pmoles

10uL

4000ng

20pmoles

20uL

We recommend 2x amount of DNA in case we need to repeat the reactions.

Protocol examples:
Example 1: If your PCR template and primer are at the concentration of 20ng/uL and 10pmol/uL, respectively, you need to add 2.5uL of PCR template and 1uL of primer to the 1.5mL epi tube, and add 6.5ul of H2O to bring the total volume of 10uL.

Example 2: If your Plasmid template and primer are at the concentration of 200ng/uL and 10pmol/uL, respectively, you need to add 2.5uL of PCR template and 1uL of primer to the 1.5mL epi tube, and add 6.5ul of H2O to bring the total volume of 10uL.

Single Tube Sequencing


Smaller orders should use 1.5mL epi tubes. Screw cap tubes are more time consuming so pop caps should be used. Each tube must be clearly labeled to match with the order forms. Tube strips while convenient are hard to label and keep track of so for large orders (24+ samples) use 96 well plates. We recommend using strip lids to cap plates as they prevent leakage and cross contamination especially for plates shipped overnight.

96 Well Plate sequencing

Please submit your DNA and primers at the concentration according to the table above. The concentration of DNA and primers should be normalized and arrayed vertically from A1-H1, A2-H2etc. on the 96 well plate. The plate should be sealed or capped using strip lids to prevent leakage and shipped overnight.

 

Pricing

 

 

Primer Extension Sequencing Service

$5 - $20 per reaction , (Inquire)

 

Direct PCR Product Sequencing Service

$5 - $20 per reaction , (Inquire)

 

Single Stranded Sequencing Service

$0.95 per base

 

Double Stranded Sequencing Service

$0.95 - $1.50 per base pair

 

Pac, Bac and P1 Ends Sequencing Service

$59 per reaction

 

cDNA/EST Library Sequencing Service

$5 - $20 per reaction (Inquire) depend on the size of the library

 

FDA Submission Sequencing Service

$1.65 - $2.45 per base pair

 

Plasmid DNAPreparation Service

$8 - $15 per sample

 

PCR Product Preparation Service

$16 - $32 per sample

 

 

 Turnaround time

 

 

Primer Extension Sequencing Service

1-2 days

 

Direct PCR Product Sequencing Service

1-2 days

 

Single Stranded Sequencing Service

1 week per 1 kb

 

Double Stranded Sequencing Service

1-2 weeks per 1 kb

 

Pac, Bac and P1 Ends Sequencing Service

2-3 days

 

cDNA/EST Library Sequencing Service

Depend on the libraries

 

FDA Submission Sequencing Service

2 weeks per 1 kb

 

Plasmid Preparation Service

1 day

 

PCR Product Preparation Service

1 day