34results about How to "Guaranteed amplification efficiency" patented technology

The invention belongs to the technical field of PCR (polymerase chain reaction), and particularly relates to a digital PCR chip with silicon substrate arrays and micro-reaction pools and a method for manufacturing the digital PCR chip. The digital PCR chip mainly comprises an upper cover and a chip. Micro-pores which are arrayed to form honeycombs are formed in the chip, and the upper cover is fixed onto a chip groove. The method includes particular steps of selecting a silicon chip with a single polished surface; cleaning the silicon chip; coating a layer of uniform photoresist on the polished surface of the silicon chip in a spinning manner; forming circular graphic arrays by means of exposure; etching silicon by the aid of a dry process in masks, namely, the circular graphic arrays to form micro-pore structures; removing the photoresist and scribing the chip to completely manufacture the digital PCR chip.

The invention belongs to the field of gene detection and especially relates to multiple PCR specific primers, kit and method for detecting a hereditary hearing loss gene, based on a high throughput sequencing technique. The primers include 52 pairs of specific primers, the nucleotide sequences of which are shown as SEQ ID NO:1 to SEQ ID NO:104. The kit further comprises the specific primers, a universal primer, an enzyme mixture, a quality control substance and nuclease-less water. The invention also discloses a detection method for capturing and amplifying the hereditary hearing loss gene through once PCR amplified reaction by utilizing the primers or kit. According to the scheme provided by the invention, the detection efficiency is effectively increased, the accuracy is increased, the cost is lowered and the operation step is simplified.

The invention discloses a multi-methylation specific PCR primer design method and system. The method comprises the steps: setting parameters according to user requirements, designing and screening primers for a target methylation site, pairing the screened multiple methylation site primer pairs in pairs, checking the compatibility, selecting a maximum number of compatible multiple primer combinations, evaluating the designed multiple primer combinations, and determining whether the primers need to be redesigned. According to the invention, multi-methylation specific PCR primer design of a super-long sequence can be realized, secondary structures such as dimer/hairpin and the like between the interiors of a single pair of primers and between multiple pairs of primers can be effectively reduced, and non-specific amplification in a genome can be effectively reduced, the designed primers are high in specificity, the operability and accuracy of the multiplex methylation specific PCR experiment testing process are improved, and the working efficiency is greatly improved.

The invention discloses sugarcane genome endogenous reference gene acetolactate synthase gene PCR (polymerase chain reaction) primer sequences and an amplification method. The primer sequences are composed of ScALS-F and ScALS-R. The PCR amplification method comprises a reaction system and amplification conditions. The invention provides a genome endogenous reference gene for PCR detection and species specific identification of transgenic sugarcane, and meanwhile, provides insurance for detection precision and result accuracy of the transgenic sugarcane.

The invention relates to a nucleic acid composition and a detection kit for detecting genetic anemia as well as a use method. The nucleic acid composition and the detection kit for detecting the genetic anemia can be used for simultaneously detecting four deletion type alpha thalassemia genes, three non-deletion type alpha thalassemia genes, nineteenth mutation type beta thalassemia genes, geneticanemia types such as sickle-shaped thalassemia gene as well as specific gene mutation types. Compared with the prior similar technologies, the detection kit for the genetic anemia has the characteristics that several mutation detection types for the genetic anemia are increased, such as alphaTHAI deleted thalassemia, sickle-shaped thalassemia and 71/72(+T) mutation and -28M(A-C) mutation which are relatively-rare thalassemia types. The detection for the genetic anemia types can provide visual reference and prompt for clinically detecting the genetic anemia, so that the leak detection risk ofclinical genetic anemia can be greatly reduced and the birth rate of severe anemia children is reduced.

The invention discloses a method for culturing and cryopreservation of CIK cells by applying a serum-free lymphocyte culture medium and obtained CIK cells. The cell culture method can develop 3*10<9> lymphocytes in a short time without introducing animal derived serum; the cell cryopreservation method can maximally improve the ratio of cryopreserved target cells and the survival rate after cell recovery. The method provides convenience for culturing, storing and related clinical research of the CIK cells.

The invention discloses qRT-PCR reference genes suitable for Rehmannia chingii Li and application of the qRT-PCR reference genes. According to the invention, six candidate reference genes with relatively stable expression quantities are screened out based on five different development stages and leaf transcriptome data of flowers of the Rehmannia chingii Li, and the stability of the candidate reference genes is analyzed through three different algorithms including GeNorm, NormFinder and Bestkeeper by utilizing a qRT-PCR technology. Results show that the number of the optimal reference genes for the flowers and leaf tissues of the Rehmannia chingii Li is two, and the optimal reference genes are RcTIP41 and Rc18S, wherein a nucleotide sequence of the RcTIP41 gene is as shown in SEQ ID NO.1,and a nucleotide sequence of the Rc18S gene is as shown in SEQ ID NO.2. According to the invention, the optimal reference genes for qRT-PCR of the flowers and leaves of the Rehmannia chingii Li are determined, and specific primers of the reference genes are provided, so that an important reference is provided for accurate and quantitative analysis of expression of related genes in the Rehmannia chingii Li.

The invention belongs to the field of gene engineering and specifically relates to endoglucanase, a coding gene coding gene cel5A-h47 and application thereof. Nucleotide sequence of the gene cel5A-h47for coding the endoglucanase is as shown in the SEQ ID No.1. Bioinformatics analysis shows that the gene-encoded endoglucanase belongs to glucoside hydrolase family 5. Through design of a primer, theendoglucanase is cloned and successfully expressed in a prokaryotic expression system; through induced ultrasonic purification, the optimum pH of the obtained recombinant endoglucanase is 5.0, and the optimum temperature is 50 DEG C. at 50 DEG C and below, relatively stable enzymatic activity can be maintained, and the endoglucanase has strong pH tolerance; and at pH 4.0-10.0, high enzymatic activity can be maintained. The recombinant endoglucanase has great research and industrial application potential.

The invention belongs to the field of gene detection and relates to multiplex PCR specific primers, kit and method for detecting causative genes of large vestibular aqueduct syndrome based on a high throughput sequencing technique. The invention discloses the multiplex PCR specific primers which are used for detecting causative genes of large vestibular aqueduct syndrome and are shown in formulas of SEQ ID NO: 1 to SEQ ID NO: 86, the kit containing the primers and the detection method for capturing and amplifying the causative gene of large vestibular aqueduct syndrome through a one-step PCR amplification reaction through the primers or kit. The multiplex PCR specific primers, kit and detection method can realizes simultaneous detection of multiple causative genes of large vestibular aqueduct syndrome of multiple samples, effectively improve the detection efficiency and accuracy, reduce a cost and simplify operation processes.

The invention belongs to the field of genetic engineering and particularly relates to an endoglucanase and its coding gene cel5A-h28 and application. The gene cel5A-h28 for encoding the endoglucanasehas a nucleotide sequence shown in the formula of SEQ ID No. 1. The bioinformatic analysis result shows that the endoglucanase coded by the gene belongs to the 5th family of glycoside hydrolase. Primers are designed and cloned and then are successfully expressed in a prokaryotic expression system. The recombinant endoglucanase obtained by induced ultrasonic purification has the optimal pH of 6.0 and the optimal temperature of 55 DEG C, keeps the stable enzyme activity at 40 DEG C, has strong pH tolerance and keeps the enzyme activity of 85% or more at pH of 4.0-9.0. The recombinant endoglucanase has the large research and industrial application potential.

The invention discloses sugarcane genome endogenous reference gene fructose-6-phosphate 2-kinase gene PCR (polymerase chain reaction) primer sequences and an amplification method. The primer sequences are composed of ScF2KP-F and ScF2KP-R. The PCR amplification method comprises a reaction system and amplification conditions. The invention provides a genome endogenous reference gene for PCR detection and species specific identification of transgenic sugarcane, and meanwhile, provides insurance for detection precision and result accuracy of the transgenic sugarcane.

The invention discloses sugarcane genome endogenous reference gene isopentenyl diphosphate delta-isomerase gene PCR (polymerase chain reaction) primer sequences and an amplification method. The primer sequences are composed of ScIDI2-F and ScIDI2-R. The PCR amplification method comprises a reaction system and amplification conditions. The invention provides a genome endogenous reference gene for PCR detection and species specific identification of transgenic sugarcane, and meanwhile, provides insurance for detection precision and result accuracy of the transgenic sugarcane.

The invention provides a kit for detecting the NUP214-ABL1 gene relative expression quantity. A real-time fluorescence PCR technology is used for detecting NUP214-ABL1 fusion gene. The kit comprises PCR reaction liquid, wherein the PCR reaction liquid comprises a probe and a primer for amplifying the NUP214-ABL1 fusion gene. Through the test, the kit has good specificity and high flexibility; the operation is simple and convenient. The detection of the NUP214-ABL1 fusion gene in a patient with acute T lymphocytic leukemia in clinics is facilitated; the important significance is realized on treatment scheme regulation, treatment effect evaluation, prognosis prediction and clinic recurrence prevention.

The invention belongs to the field of genetic engineering and particularly relates to an endoglucanase and its coding gene cel5A-h55 and application. The gene cel5A-h55 for encoding the endoglucanasehas a nucleotide sequence shown in the formula of SEQ ID No. 1. The bioinformatic analysis result shows that the endoglucanase coded by the gene belongs to the 5th family of glycoside hydrolase. Primers are designed and cloned and then are successfully expressed in a prokaryotic expression system. The recombinant endoglucanase obtained by induced ultrasonic purification has the optimal pH of 5.0 and the optimal temperature of 50 DEG C, keeps the stable enzyme activity at 50 DEG C or less, has strong pH tolerance at pH of 5.0-9.0 and keeps the high enzyme activity. The recombinant endoglucanasehas the large research and industrial application potential.