Indian Council of Agricultural Research
(Agricultural Knowledge Management Information System)


ICAR-Indian Grassland & Fodder Research Institute, Near Pahuj Dam, Gwalior Road, Jhansi-284 003 Uttar Pradesh (www.igfri.res.in)
Click Here to See Management Committee
Instrument Details
Instrument/Facility Name: Real Time PCR
Make                                   : Eppendorf
Model                                  : Master Cycler ep realplex
Specification   :

Click to ViewThe Eppendorf Mastercycler® RealPlex2 is a small, robustly designed instrument for real-time PCR. The machine features a special 96-well heating block, which enables rapid thermal cycling. The RealPlex2 contains a maximum of two filters, which enables fluorescence monitoring at 520 and 550nm. Hence the machine is compatible with the most widely used reporting dyes (SYBR Green, FAM, JOE & VIC).

  Working Principles
Click to ViewReal-Time PCR Thermal Cyclers/ Thermo cyclers, also referred to as Quantitative PCR (qPCR) calculate PCR amplification as it occurs. Using smaller amounts of starting material and combining nucleic acid amplification and detection allows for efficiency and eliminates the post-amplification process. Real-time PCR thermo cyclers can be used with both DNA and RNA.Synthesis of cDNA from total RNA samples is the first step in the two-step RT-PCR gene expression quantification experiment. Real-time or Quantitative PCR and RT-PCR use the linearity of DNA amplification to determine absolute or relative amounts of a known sequence in a sample. By using a fluorescent reporter in the reaction, it is possible to measure DNA generation.In quantitative PCR, DNA amplification is monitored at each cycle of PCR. When the DNA is in the log linear phase of amplification, the amount of fluorescence increases above the background. The point at which the fluorescence becomes measurable is called the Quantification Cycle (Cq) or crossing point. By using multiple dilutions of a known amount of standard DNA, a standard curve can be generated of log concentration against Cq. The amount of DNA or cDNA in an unknown sample can then be calculated from its Cq value.

  Applications
  User Instructions
Real-time PCR is used for sensitive, specific detection and quantification of nucleic acid targets. Applications in many scientific areas of molecular biology for gene expression analysis, SNP genotyping, micro RNA and non-coding RNA analysis# Use powder-free gloves and RNase free H20 while preparing the samples # To avoid high amount of bound dye to initial template use not more than 50 ng of DNA or 100ng of RNA in total volume of 20 ul. # Protect SYBR green from light and avoid repeated freezing and thawing # Final concentration of the primer and probe should be 300-900nM

  Contact Us
Contact No. 05102730666
Email Id igfri.director@gmail.com; director.igfri@icar.gov.

  Real Time PCR Charges
Procedure Name
Industry
University
National LAB/R&D
Remark
2500/ reac
2000/ reac
2000/ reac

Additional Rs 100/-towards postal charges if result are to be posted
The Demand Draft should be in favour of ""
Letter, DD & Samples send to "Letter and sample should be sent to "The Director, IGFRI, Jhansi-284 003""