The PCR analysis was performed using 10 μL reaction mixtures containing 20 ng of total genomic DNA, 2 pM of primer, and 5 μL of GoTaq Green Master Mix (Promega, madison, WI, USA). PCR was performed under conditions of 95°C for 5 min and subsequent 35 rounds of 94°C for 30 s, 45°C for 30 s, and 72°C for 30 s, using a Biometra T1 Thermocycler (Biometra, Goettingen, Germany). The PCR products were separated by electrophoresis in 3% gel of certified low range ultra-agarose (Bio-rad) followed by ethidium bromide staining.
Biometra t1 thermocycler
Manufactured by Analytik Jena
Sourced in Germany
The Biometra T1 Thermocycler is a laboratory instrument designed for the amplification of DNA or RNA samples. It is a compact, programmable thermal cycler capable of performing the temperature cycling required for various molecular biology techniques, such as polymerase chain reaction (PCR).
Automatically generated - may contain errors
3 protocols using biometra t1 thermocycler
1
PCR Analysis of Genomic DNA
2
MLPA Probe Mix Analysis for Copy Number
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Two SALSA MLPA probe mixes were used: P027 (version C1, lot 0213; MRC-Holland) containing 50 probes and P105 (version D1, lot 0413) containing 55 probes. These two probe mixes were selected for the MLPA analysis as both contain probes targeting multiple genomic regions on chromosomes 1 to 20 and 22. For details about the probe mixes, see Supplementary Tables S1 and S2 (all supplemental materials can be found at American Journal of Clinical Pathology online). Commercially obtained blood-derived genomic DNA of multiple anonymous male or female donors (Human Genomic DNA: Male [G1471] and Female [G1521]; Promega, Mannheim, Germany) was used as a normal diploid copy number reference sample DNA for normalization. All MLPA reactions were performed using a standard MLPA one-tube protocol (version MDP-005, available at www.mlpa.com ) on a Biometra T1 Thermocycler (Biometra, Goettingen, Germany). Fragment separation was performed on an ABI PRISM 3100xl Genetic Analyser (4359571; Life Technologies). The generated raw data were analyzed with Coffalyser.Net v.140721.1958 software. Intrasample normalization and intersample normalization were done using a population analysis method. Copy number ratio values between 0.8 and 1.2 were considered normal.
3
Characterization of DRD4 Exon III Repeat
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DNA was extracted by Master Pure kit (Epicentre, Madison WI) and PCR amplification was carried out. The exon III repeat region of the DRD4 receptor was characterized by a PCR amplification procedure (using a Reddy Mix kit, AB gene, Surrey UK) with the following primers: F5′-TTCCTACCCTGCCCGCTCATGCTGCTGCTCATCTGG-3′; R5′-ACCACCACCGGCAGGACCCTCATGGCCTTGCGCTC-3′. We performed PCR reactions using 5 μl Master Mix (Thermo scientific), 2 μl primers (0.5 μM), 0.6 μl Mg/Cl2 (2.5 mM), 0.4 μl DMSO 5%, and 1 μl of water to total 9 μl volume; an additional 1 μl of genomic DNA was added to the mixture. All PCRs were employed on a Biometra T1 Thermocycler (Biometra, Güttingem, Germany).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!