46results about How to "Reduce the starting amount" patented technology

The invention discloses a method for constructing an absolute quantitative transcriptome library based on a unique recognition sequence. Using the fragmented mRNA as a template, the first cDNA strand is synthesized under the action of reverse transcriptase using a primer pool with a universal linker sequence, and a unique recognition UID sequence is added to the 3' end of the synthesized cDNA by an enzymatic reaction. Build a library linker so that each cDNA has a unique sequence tag; finally, use a universal library linker to perform PCR amplification to obtain an RNA library. For the first time, the present invention uses the splint ligation method to construct an RNA library based on single-stranded cDNA. At the same time, the UID sequence is used to accurately restore the cDNA composition before PCR amplification, which can realize accurate quantification of transcripts; the present invention uses single-stranded cDNA as a raw material for library construction. The step of second-strand synthesis is omitted, the template loss rate is reduced, cost and time are saved, and the existing technology can only relatively quantify transcripts.

The invention relates to a breast cancer susceptibility gene variable library construction method. The construction method comprises the following steps that 1, according to variation regions of a breast cancer-related gene, primer pairs capable of detecting the variation regions are designed; 2, judgment parameters R of the primer pairs are calculated, the primer pairs of which values of the judgment parameters are smaller than or equal to 1 are classified into a first primer combination solution, and the primer pairs of which values of the judgment parameters are larger than 1 are classified into a second primer combination solution; 3, a to-be-detected DNA sample is extracted and purified; 4, initial PCR amplification is conducted on the purified sample by adopting the first primer combination solution and the second primer combination solution separately; 5, linker connection is conducted on a target fragment to obtain a fragment with a linker; 6, library PCR amplification is conducted through a mixed solution of the first primer combination solution and the second primer combination solution to obtain a sequencing library. The library constructed through the breast cancer susceptibility gene variable library construction method is high in sequencing flux, sensitivity and specificity and can be used for detecting low-frequency mutation of free DNA.

The invention belongs to the technical field of genetic engineering, and in particular relates to a single-cell chromosome aneuploidy detection system and its application. The detection system includes a window division module, a segment division module, a segment copy rate calculation module, and a segment aneuploidy judgment module. The detection system uses up to 700k reads / sample, and can detect chromosome deletion / duplication of more than 5Mb in the sample, with an accuracy rate of up to 99.9%, reducing the depth of sequencing and reducing costs.

The invention relates to a colorectal cancer susceptibility gene mutation library constructing method. The colorectal cancer susceptibility gene mutation library constructing method comprises the following steps: A, designing primer pairs capable of detecting a mutation region according to the mutation region related to a colorectal cancer; B, calculating to obtain judgment parameters R of the various primer pairs, classifying the primer pairs of which the values of the judgment parameters R are smaller than or equal to 1 into a first group of primer combination liquid, and classifying the primer pairs of which the values of the judgment parameters R are greater than 1 into a second group of primer combination liquid; C, extracting and purifying DNA of a sample to be detected; D, carrying out initial PCR amplification on the purified sample by using the first group of primer combination liquid and the second group of primer combination liquid respectively; E, connecting target fragments via joints to obtain fragments with joints; and F, carrying out library PCR amplification by using mixed liquid of the first group of primer combination liquid and the second group of primer combination liquid to obtain a sequencing library. The library constructed by the colorectal cancer susceptibility gene mutation library constructing method is high in sequencing flux, high in sensitivity and high in specificity, and can be used for detecting free DNA low frequency mutation.