Do you perform DNA extraction?

Yes, we offer DNA extraction service for isolating environmental DNA from any sample type. We use commercial DNA isolation kits from QIAGEN and automate the process of DNA extraction using a QIACube machine. We store leftover samples for three months, after which the samples will be discarded.

Which sample types are accepted?

We accept all types of samples, e.g., soil, sediment, sludge, manure, stool, meat, tissue, swabs, river, lake, wastewater, and seawater.

What are the required volume and concentration of the liquid samples?

We require 1-10 L of water for river, lake, and seawater samples. For wastewater, we require 100 mL from influent and 250 mL from effluent. The sample should be filtered/concentrated using a hydrophilic membrane filter with a minimum pore size of 0.2 µm. We recommend using PES membrane. Each concentrated water-filter should be stored in an individual 50 mL-falcon tube and delivered on dry ice to our facility in Helsinki, Finland.

What are the required volume and concentration of the solid samples?

For solid samples, we recommend a dry/wet weight of 100 g. The solid samples should be stored in a zip lock plastic bag or a safe-lock 1.5 mL microtube and delivered on dry ice to our facility in Helsinki, Finland.

Our DNA samples are ready, what is the required volume and concentration of DNA samples?

For chip analysis of over 100 genes: using NanoDrop One measurement, we recommend 10 ng/uL and 100 uL of volume, with DNA quality of 260/280 between 1.8-2.0 +/- 0.1. For chip analysis of less than 100 genes: the minimum volume of samples is 50 uL. If you have samples with low amounts of DNA, the minimum DNA concentration is 3 ng/uL. Optimal sample concentrations help us ensure that you get the best results.

We have the DNA samples, but they are not diluted. Can we send our DNA samples without diluting them to the required concentration?

Yes, you can still send us your DNA samples. However, there will be an extra dilution service fee.

Which genes do you measure? Can we choose the target genes?

In each chip analysis, we can measure up to 384 genes, including 16S rRNA gene (positive control); antibiotic resistance genes (ARGs) targeting aminoglycoside, amphenicol, beta lactam, florphenicol, multidrug efflux-pump, macrolide-lyncoside-streptoganin B (MLSB), trimethoprim, tetracycline, vancomycine; genes targeting other antibacterials such as nisin, bacitracin, antispetic, mercury; and genes that are associated with mobile genetic elements (MGEs) and integrons.

Yes, you can customise the target genes with an extra customisation fee. The customisation options for chip analysis are: 296 Genes: 5 Samples; 248 Genes: 6 Samples; 216 Genes: 8 Samples; 144 Genes: 12 Samples; 120 Genes: 14 Samples; 96 Genes: 18 Samples; 80 Genes: 21 Samples; 72 Genes: 24 Samples; 54 Genes: 32 Samples; 48 Genes: 36 Samples; 36 Genes: 48 Samples; 24 Genes: 72 Samples; and 12 Genes: 128 Samples. You can choose the target genes based on over 600 previously validated primer sets that are available in our database from the the ARG selection sheet that we will send you separately.

How are the genes measured?

The ARG 1.0 and ARG 2.0 primer sets were developed to detect and quantify up to 384 genes in parallel and in one measurement using the SmartChip Real-Time PCR system. We updated the 384 gene ARG 2.0 primer sets to the ARG 2.1 primer sets according to the current antibiotic resistance gene database (CARD) to mantain the specificity of primer sets.

How do you account for variation in the genes in the primer design?

Each primer set was designed to target a specific region in an antibiotic resistance gene (ARG). For ARG variations, more primers have been designed to cover all known ARG variations.

Do you detect pathogens?

No, at the moment we only detect antibioitic resistance genes. However, eight primer sets are available to target certain taxonomy group of bacteria: A. baumannii, Bacteroidetes, Campylobacter, Enterococci, Firmicutes , K. pneumoniae, P. aeruginosa, and Staphylococci.

Are the results binary or quantitative?

We use the delta Ct calculation. Delta Ct = Ct of genes – Ct of 16S rRNA gene. The relative quantification is calculated with formula: 2^-delta Ct.

How long is the analysis process?

We deliver analysis results within 10 working days. For three or more chips, the analysis process may take longer.

Can you add custom targets to the SmartChip qPCR measurement?

Yes, if the primer sets are already validated, have annealing temperatures of 60 °C in quantitative PCR, and a positive control of the gene is available.

How quickly do you add new genes to your offering?

It takes at least 1-3 months to optimise and validate new genes. Providing us the positive control of the new gene will make the validation process faster.

Do you provide analysis of the SmartChip qPCR measurement results?

Yes, we can analyse raw data from the SmartChip qPCR measurement to provide meaningful gene abundances in each sample. We deliver the analysis results via ResistApp, an interactive online dashboard, which displays the numbers, abundances, and copy numbers of detected genes relative to the 16S rRNA genes, as well as a heatmap of gene profiles.

Do you provide further or custom analysis of the SmartChip qPCR measurement results?

Yes, we provide statistical analyses such as ordination, fold change, and gene network analysis as an additional service.

How to prepare the DNA samples to be delivered to Resistomap?

For DNA samples that include up to 25 samples, we recommend using safe-lock 1,5 mL microtubes to avoid evaporation or spilling during transportation. Please label the tubes individually in numerical order only, e.g., 1-25. DO NOT USE PCR tubes for sending your samples. For DNA samples that include over 25 samples, we recommend using a sealed 96-well plate. Please add the samples according to the following template per chip analysis. Finally, please provide information about your samples in the Sample specification sheet, which we will send you before you send the samples.

How to send samples to Resistomap?

Please send the samples inside a protected box at room temperature to our facility: Resistomap Oy, Viikinkaari 4C, Helsinki 00790, Finland. Contact: Jesse Majlander, Phone: +358 500500755. Please email the tracking number to: Jesse@resistomap.com. We recommend using our courier partner, DHL Express. UK customers should use the following EORI No: FI2921012-4.

Can we get back the leftover DNA samples after the analysis?

No, usually there are only a few uL leftover from DNA samples after the SmartChip qPCR analysis. However, if you order the DNA extraction service at our facility, we can send back the leftover DNA extraction samples.

What is the final concentration of DNA sample that is analysed in each qPCR reaction in the SmartChip qPCR measurement?

In each 100 uL of qPCR reaction, a total of 2 ng/uL of DNA sample is measured.

When we choose the SmartChip qPCR service with data analysis, the results are delivered via ResistApp. What is ResistApp?

ResistApp is an online intearctive dashboard, which displays the numbers, abundances, and copy numbers of detected genes relative to the 16S rRNA genes, as well as a heatmap of gene profiles. You can filter the results according to your interest e.g., view gene abundances for a certain group of antibiotics or view results by sample type by clicking the shift button. With ResistApp, you can share and present the results effciently to your colleagues by giving them access to your project.

How are the gene copy numbers in ResistApp calculated? Are the results not relative to 16S rRNA genes?

The results from SmartChip qPCR measurement are relative abundances to 16S rRNA genes. In ResistApp, the gene copy numbers are estimated gene abundances against known copy numbers of 16S rRNA genes in each sample. To calculate copy numbers of 16S rRNA genes in each sample, we measure the standard curve for 16S rRNA genes in the SmartChip qPCR measurement.

Can we measure the copy numbers of target genes?

Yes, you can measure the gene copy numbers by providing at least four to five serially diluted samples as the standard curve of the target gene. The standard curve samples will be measured with the other samples in the same chip analysis. You can calculate the target gene copy numbers based on the Ct values of the diluted samples used as the standard curve.