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Beijing, China

Patent
Berry Genomics Co. | Date: 2012-01-10

The disclosure relates a method and a kit for constructing a plasma Deoxyribonucleic acid (DNA) sequencing library. The method provided by the disclosure includes: extracting a plasma DNA; making the plasma DNA ligate to a sequencing linker, and purifying a ligation product; performing Polymerase Chain Reaction (PCR) amplification for the purified ligation product, purifying the PCR amplification product, and obtaining the plasma DNA sequencing library, wherein, the method does not include the step of performing 5-terminus phosphorylation for the plasma DNA. The kit provided by the disclosure includes: a reagent which ligates a plasma DNA to a sequencing linker, including the sequencing linker, a ligase and a ligation buffer; and reagents and instruments for purifying the ligation product; a reagent which performs PCR amplification for a purified ligation product, and reagents and instruments for purifying the PCR amplification product; wherein, the kit does not include the reagent which performs 5-terminus phosphorylation for the plasma DNA. The disclosure simplifies the construction flow of a plasma DNA sequencing library, simplifies the experimental procedures, and makes the construction of the plasma sample library have lower cost, higher efficiency and faster speed, and is convenient for large-scale application.


The disclosure claims a cleaved Deoxyribonucleic acid (DNA) detection method, a DNA fragment detection kit and use thereof. Wherein, the method includes the steps of: designing primers according to a test site or a test region of the DNA fragment; cyclizing the DNA fragment to obtain acyclized DNA; implementing Polymerase Chain Reaction (PCR) amplification for the cyclized DNA by using the primers; and detecting the PCR amplification product. In the disclosure, by cyclizing the DNA fragment, the amplification can be implemented even if only one PCR primer can match with a template, thus, the adaption range and effective template amount of the primer amplification can be greatly increased, and the detection sensitivity of the DNA fragment can be greatly improved.


The present invention is directed to a method, kit and primers for detecting fetal deafness pathogenic gene mutations. The method of the invention comprises: (a) designing primers according to the pre-determined mutation loci of deafness pathogenic genes; (b) extracting plasma DNAs in a pregnant woman; (c) connecting the extracted plasma DNAs with pre-amplification linkers to obtain connected products; (d) PCR pre-amplifying the connected product to obtain pre-amplified products; (e) cyclizing the pre-amplified products to obtain cyclised DNAs; (f) PCR amplifying the cyclised DNAs using the designed primers to obtain amplified products; and (g) high throughput sequencing the amplified products and analyzing the mutations of the fetal deafness pathogenic genes. The invention can effectively determine whether the pre-determined loci on deafness pathogenic genes have been mutated as well as the mutation type.


The disclosure claims a method for tracking a sample in a second-generation Deoxyribonucleic acid (DNA) sequencing technology and a detection kit, wherein the method includes the following steps of: 1) incorporating DNA molecular tag with a known sequence into a sample, and obtaining a sequencing sample; 2) sequencing the sequencing sample; 3) screening the molecular tag sequence from the sequencing result of step 2), and comparing with the known sequence of the molecular tag. As the sequencing process of the tag is synchronously implemented during the sequencing process of the DNA molecular, this method can be conveniently operated, and the confusion of the samples caused by manual operation can be found instantly; thereby, this method not only has important significance for the technical research, but also greatly improves the strictness of the clinical detection if applied to the clinical detection.


Shaw S.W.S.,Chang Gung University | Hsiao C.-H.,Taipei Medical University Hospital | Hsiao C.-H.,National Yang Ming University | Chen C.-Y.,Taipei Veterans General Hospital | And 5 more authors.
Fetal Diagnosis and Therapy | Year: 2014

Objective: To evaluate the performance of noninvasive prenatal testing for all fetal chromosomal aneuploidies in an extremely high-risk group undergoing first trimester combined Down syndrome screening. Method: A multicenter cohort prospective study in Taiwan was performed between June and December 2012. Maternal plasma was collected and shotgun massive parallel sequencing was performed on each fetal chromosome. 201 Taiwanese pregnant women at >12 weeks' gestation from 11 medical centers were enrolled in this trial. The extremely high-risk group was defined as a Down syndrome risk cutoff >1:30 or nuchal translucency >3.0 mm (n = 100), while the low-risk group was defined as a Down syndrome cutoff <1:1,500 (n = 101). Amniocentesis confirmation was performed and birth outcome was also recorded. Results: There were 11 cases of trisomy 21, 8 cases of trisomy 18, 3 cases of trisomy 13, 1 case of trisomy 16, 3 cases of 45,X, and 1 case of 47,XYY detected prenatally in 100 extremely high-risk gravidas [n = 27/100 (27%)]. The overall autosomal or sex chromosome aneuploidy detection rate was 96% (27/28) because of an insufficient amount maternal plasma for one fetus with Turner syndrome. In the low-risk group, no chromosomal abnormalities were detected (specificity = 100%). There were no false-positive cases in this study. Conclusions: This first trial in Taiwan shows that noninvasive prenatal testing for whole chromosome aneuploidies can be efficiently applied in extremely high- and low-risk populations. © 2013 S. Karger AG, Basel. Source

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