Frequently Asked Questions

What is Laragen's policy for repeat sequences?

We will provide free repeats to troubleshoot failed reactions. If a large number of reactions failed, we will repeat a few samples to troubleshoot. If the repeated reactions work, we can repeat all failed reactions at a charge upon request.

Why didn't my sequence work?

  • Poor quality of template DNA: Contaminants in the DNA prep will interfere with the reaction and cause failures, noise and miscalls. Contaminants are ethanol, protein, salt, PEG, EDTA and genomic DNA. We recommend Qiaprep Spin Column miniprep kit to prepare plasmid DNA. Do not overgrow the cultures and do not overload the spin column.

  • Not enough DNA: Insufficient DNA will lead to insufficient signals or failed reactions. On the other side, too much DNA will also interfere with the reaction.

  • EDTA inhibits sequencing reactions, Do not elute DNA with TE buffer. Elute with water or EB(10mM TrisHCL, PH8.0)

  • Primer concentration: 10 pmole of primer is required per sequencing reaction. Make sure that your primer concentration is correct. Too much or too little primer may result in sequencing failures. Based on our experience, you should test your sequencing primer as a PCR primer first. If the primer works as a PCR primer, it usually works fine as a sequencing primer. However, since PCR reactions is more robust than sequencing reaction, some primer may work well for PCR reactions, but not for sequencing reactions.

  • Difficult Templates: Some templates are difficult to sequence. For example, it is difficult to sequence a GC rich region with regular BigDye premix. If that is the case, we will use Laragen's proprietary chemistry for difficult templates to get around it. Difficult templates include sequences with GC rich region, hairpin, shRNA and RNAi etc.

How can I retrieve my sequences?

After your sequences are ready, you will receive a notification email. You can login into your account here to download your sequences.

How many base pairs can I get per read?

It depends on the quality of the DNA templates and the sequences. You can get > 900 bp per read on average. If you want to get more than 900 bp, please contact us and we will reanalyze the sequence for you.

How close can I read from the primer?

It depends on the quality of the DNA templates. You can get reliable sequences starting from 17 - 25 bp away from the primer.

How and where can I ship my samples to Laragen?

You can ship your DNA sample either in water or EB buffer (No EDTA) by US mail or FedEx (preferred) at room temperature to:

Laragen, Inc
10601 Virginia Ave
Culver City, CA 90232

We also provide free pick-up in the Los Angeles Metro area.

What is the turnaround time?

Type of sample Turnaround Time
Sequencing ready-to-run sample Overnight
Sequencing premix sample Overnight
Sequencing purified plasmid and PCR sample Overnight
Sequencing un-purified PCR sample Overnight
Bacterial cell (RCA) 24 to 48 hours
Genotyping animal tissue with gel-based or qPCR 48-72 hours
Genotyping animal gDNA with gel-based or qPCR 48-72 hours
Fragment ready-to-run analysis Overnight
Microbial ID coloines or cell pellet 5 to 7 days
Genotyping copy number analysis 72 hours
Cell line authentication 3-5 days

What primers do you provide in-house?

Standard Primers Sequences
M13 Forward GTA AAA CGA CGG CCA GT
M13 Reverse CAG GAA ACA GCT ATG AC
T7 Promoter TAA TAC GAC TCA CTA TAG GG
T3 ATT AAC CCT CAC TAA AG
SP6 ATT TAG GTG ACA CTA TAG
T7 Terminator CTA GTT ATT GCT CAG CGG TG
pGEX 5' GGG CTG GCA AGC CAC GTT TGG TG
pGEX 3' CCG GGA GCT GCA TGT GTC AGA GG
RV3 CTA GCA AAA TAG GCT GTC CCC
BGH Reverse TAG AAG GCA CAG TCG AGG C
pBADfor ATG CCA TAG CTT TTT ATC C
pBADrev GAT TTA ATC TGT ATC AGG
pFastBacFwd TAT TCC GGA TTA TTC ATA CCG TC
pFastBacRev GTA TGG CTG ATT ATG ATC CTC
CMVfor CGC AAA TGG GCG GTA GGC GTG
CMVRev AGT AGG AAA GTC CCG TAA GG

How long do you keep my DNA samples and primers?

Unless you specifically request, we normally keep our clients' DNA samples and primers for three weeks. The DNA samples and primers will be trashed after three weeks.

Why is the DNA concentration you measured lower than measured by nanodrop?

We use a fluorometer coupled with Hoechst 33342 dye to measure DNA concentration. Because Hoechst 33342 only binds to double strand DNA, not protein, RNA or other contaminants, the DNA concentration measured with this technique is more accurately measured. In comparison, nanodrop is based on absorption peak at 260/280 nm wavelength. Many contaminants in the DNA solution could affect the OD reading. We found if the DNA concentration measured by fluorometer is significantly lower than the client's concentration measured by spectrophotometer (such as nanodrop) the sequencing reactions tends to fail due to the contaminants present in the DNA samples.

How Can I view AB trace files

Sequence Scanner is a powerful software to view AB trace files. Sequencing Scanner is able to provide a variety of QC data and is able to view multiple trace files.

Chromas 2.6.4 is a PC based freeware to view AB trace files.

4Peaks V1.8 is a Mac OSX 10.7 or above based freeware to view AB trace files.

Download Service Request Forms

Sequencing Template

Genotyping Request Form

MiSeq Request Form

qPCR Ready-To-Run Request Form

Fragment Analysis Request Form

SNP Genotyping Request Form

Cell ID Shipping Instruction

Cell ID Service Request Form

Microbial ID Request Form

Trace file sample to demonstrate long sequencing reads

Instructions for Downloading Your Sequencing Results

All new users will need to create a login account on our DNALIMS server here. For other services, please email a copy of the corresponding request form.

Sequencing clients will receive an automated email notification when the sequences are ready to download. You can enter more than one email address in the email field separated by commas.

After you receive the email notification, you can login to your DNALIMS account and download your sequences

For all other services, you will receive emails with your results attached in Excel format. The raw data, such as .fsa files, are available to download from dropbox link we send.

We recommend you use either Sequence Scanner or Chromas 2.6.4(PC-Based) and 4Peaks V1.8(OSX 10.7+) to view electropherograph file (.ab1). We set our sequencing read length at 1000 bp. If the sequences are good and you would like to have longer reads, you can submit the .ab1 files to PeakTrace.

Laragen, Inc
10601 Virginia Ave, Culver City, CA90232
310-280-0804 info@laragen.com