The table included below shows recommended and maximal input amounts for the various sample types that can be processed with the Monarch Genomic DNA Purification Kit (NEB #T3010). Additionally, typical yields and DIN values are shown. Using input amounts that exceed the recommended amount will lead to a reduction of yield and purity in those samples. If more starting material is required, splitting the sample and processing on multiple columns is recommended. For low input amounts where expected yields are <100 ng (which corresponds to 5 µl whole blood, 1 x 104 cultured cells or 0.2 mg tissue), the use of carrier RNA is recommended (See “Using Carrier RNA for Low Inputs” in the product manual).
The demands on the purification process for tissue samples vary highly between different sample types. Therefore, different input amounts are indicated for individual groups of tissues and additional guidelines are provided in the protocol ensuring that the best possible results are obtained for each tissue type.
|SAMPLE TYPE||RECOMMENDED INPUT AMOUNT||TYPICAL YIELD (μg)||DIN||MAXIMUM INPUT AMOUNT|
|Tail (mouse)||10 mg||12–20||8.5–9.5||25 mg|
|Ear (mouse)||10 mg||18–21||8.5–9.5||10 mg|
|Liver (mouse and rat)||10 mg||15–30||8.5–9.5||15 mg|
|Kidney (mouse)||10 mg||10–25||8.5–9.5||10 mg|
|Spleen (mouse)||10 mg||30–70||8.5–9.5||10 mg|
|Heart (mouse)||10 mg||9–10||8.5–9.5||25 mg|
|Lung (mouse)||10 mg||14–20||8.5–9.5||15 mg|
|Brain (mouse and rat)||10 mg||4–10||8.5–9.5||12 mg|
|Muscle (mouse and rat)||10 mg||4–7||8.5–9.5||25 mg|
|Muscle (deer)||10 mg||5||8.5–9.5||25 mg|
|Human (whole)||100 μl||2.5–4||8.5–9.5||100 μl|
|Mouse||100 μl||1–3||8.5–9.5||100 μl|
|Rabbit||100 μl||3–4||8.5–9.5||100 μl|
|Pig||100 μl||3.5–5||8.5–9.5||100 μl|
|Guinea pig||100 μl||3–8||8.5–9.5||100 μl|
|Cow||100 μl||2–3||8.5–9.5||100 μl|
|Horse||100 μl||4–7||8.5–9.5||100 μl|
|Dog||100 μl||2–4||8.5–9.5||100 μl|
|Chicken (nucleated)||10 μl||30–45||8.5–9.5||10 μl|
|HeLa||1x106 cells||7–9||9.0–9.5||5x106 cells|
|HEK293||1x106 cells||7–9||9.0–9.5||5x106 cells|
|NIH3T3||1x106 cells||6–7.5||9.0–9.5||5x106 cells|
|E. coli (Gram-negative)||2 x 109 cells||6–10||8.5–9.0||2 x 109 cells|
|Rhodobacter sp. (Gram-negative)||2 x 109 cells||6–10||8.5–9.0||2 x 109 cells|
|B. cereus (Gram-positive)||2 x 109 cells||6–9||8.5–9.0||2 x 109 cells|
|T. kodakarensis||2 x 109 cells||3–5||8.5–9.0||2 x 109 cells|
|S. cerevisiae||5 x 107 cells||0.5–0.6||8.5–9.0||5 x 107 cells|
|Saliva (human)||200 µl||2–3||7.0–8.0||500 µl|
|Buccal swab (human)||1 swab||5–7||6.0–7.0||1 swab|
* Tissue gDNA yields are shown for frozen tissue powder, frozen tissue pieces and RNAlater-stabilized tissue pieces. Though frozen tissue powder results in highly-intact gDNA, lower yields can be expected than when using frozen or RNAlater-stabilized tissue pieces. Residual nuclease activity in tissue pieces will cut the gDNA, resulting in a slightly smaller overall size; however, this gDNA is optimal for silica-based purification.
** Human whole blood samples stabilized with various anticoagulants (e.g. EDTA, citrate and heparin) and various counter-ions were evaluated and results were comparable in all cases. Additionally, all indicated blood samples were tested both as fresh and frozen samples, yielding comparable results. Human samples were donated by healthy individuals; yields from unhealthy donors may differ.
*** Buccal swabs and saliva samples partially consist of dead cell material with degraded gDNA. Therefore, the purified gDNA from those samples will naturally have lower DIN values.
Additional Resources you may find helpful:
- Monarch Genomic DNA Purification Kit Product Manual
- Troubleshooting Guide for Genomic DNA Extraction & Purification
- Factors Affecting DNA Quality when Purifying gDNA from Blood and Tissues with the Monarch Genomic DNA Purification Kit
- Guidelines for Handling Tissue Samples when using the Monarch Genomic DNA Purification Kit