Highly multiplex PCR methods and compositions

A technology of reaction mixtures, primer pairs, applied in the field of highly complex PCR and composition, which can solve the problems of invasive procedures, poor accuracy, low accuracy, etc.

Inactive Publication Date: 2015-06-03
NATERA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Unfortunately, standard methods are either inaccurate or involve invasive procedures that risk miscarriage
Methods based on maternal blood hormone levels or ultrasound measurements are non-invasive, but they are also less accurate
Methods such as amniocentesis, chorionic villi biopsy, and fetal blood sampling are highly accurate, but they are invasive and carry significant risks
In the United States, amniocentesis is performed in approximately 3% of all pregnancies, but its use has declined in frequency over the past fifteen years

Method used

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  • Highly multiplex PCR methods and compositions
  • Highly multiplex PCR methods and compositions
  • Highly multiplex PCR methods and compositions

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Embodiment Construction

[0181] The present invention is based in part on the unexpected discovery that generally only a relatively small number of primers in a primer library results in a large number of amplified primer dimers formed during the multiplex PCR reaction. Methods have been developed to select the least desirable primers for removal from the library of candidate primers. By reducing the amount of primer dimer to a negligible amount (about 0.1% of the PCR product), these methods allow the resulting primer library to simultaneously amplify a large number of target loci in a single multiplex PCR reaction. Because the primer hybridizes to the target locus and amplifies it instead of hybridizing to other primers and forming an amplified primer dimer, the number of different target loci that can be amplified increases. It has also been found that using a lower than normal primer concentration and a much longer annealing time increases the likelihood that the primers will hybridize to the target...

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Abstract

The invention provides methods for simultaneously amplifying multiple nucleic acid regions of interest in one reaction volume as well as methods for selecting a library of primers for use in such amplification methods. The invention also provides library of primers with desirable characteristics, such as minimal formation of amplified primer dimers or other non-target amplicons.

Description

[0001] Cross references to related applications [0002] This application claims the rights and priority of U.S. Utility Application No. 13 / 683,604 filed on November 21, 2012 and U.S. Provisional Application No. 61 / 675,020 filed on July 24, 2012. U.S. Utility Application No. 13 / 683,604 is a partial continuation application of U.S. Utility Application No. 13 / 300,235 filed on November 18, 2011, and a partial continuation of U.S. Utility Application No. 13 / 110,685 filed on May 18, 2011 Apply and claim the rights of U.S. Provisional Application No. 61 / 675,020 filed on July 24, 2012. U.S. Utility Application No. 13 / 110,685 requires U.S. Provisional Application No. 61 / 395,850 filed on May 18, 2010; U.S. Provisional Application No. 61 / 398,159 filed on June 21, 2010; Filed on February 9, 2011 United States Provisional Application No. 61 / 462,972; United States Provisional Application No. 61 / 448,547 filed on March 2, 2011; and United States Provisional Application No. 61 / 516,996 filed on A...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G16B20/10G16B20/20G16B30/00
CPCC12Q1/686G16B20/00G16B30/00G16B20/20G16B20/10C12Q2537/143G16B25/20
Inventor B·齐默曼M·M·希尔P·G·拉格劳特M·多德
Owner NATERA
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