45 results about "Amplified fragment length polymorphism" patented technology

The invention provides a fluorescently-labeled gene multiplex amplification technology for detecting sources of goat, sheep, pig and duck meat simultaneously. A method comprises the steps that mitochondria 16S rRNA (ribosomal Robonucleic Acid) and Cyt (Cytosine) b genes of a goat, sheep, a pig and a duck serve as target genes, and are subjected to PCR (Polymerase Chain Reaction) multiplex amplification by two pairs of fluorescently-labeled primers; PCR amplification products are detected by a 3730xl full automatic gene analysis meter; and species identification is performed according to 16S rRNA gene amplified fragment length polymorphism. The method fills in a gap of a detection method of the four kinds of meat.

The invention belongs to the field of inheritance breeding, and relates to a molecular marker for a single fruit weight main-effect quantitative trait loci (QTL) of Dangshansu pear fruit and application thereof. A genetic linkage map of Dangshansu pear is constructed by utilizing sequence-related amplified polymorphism (SRAP), simple sequence repeats (SSR) and amplified fragment length polymorphism (AFLP), phenotypic identification on the single fruit weight of the fruit is combined, a group is analyzed through interval mapping, the presence of the main-effect QTL is detected on the 13th linkage group of the Dangshansu pear, and the 48.8 percent of single fruit weight can be interpreted. A molecule closest to the main-effect QTL Pfw-1 is marked as fe3em4-242p* and the distance is 0cM. The obtained molecular marker for the single fruit weight main-effect QTL has important theoretical and practical guidance significance for quickening the genetic improvement progress of pear varieties and improving breeding selection efficiency.

The invention provides a striped beakfish sex specific molecular marker and a genetic sex identification method thereof. The method comprises: screening and cloning of a male specific amplified fragment length polymorphism (AFLP) marker; performing DNA sequence analysis on sex specificity; and constructing a genetic sex polymerase chain reaction (PCR) identification method. In the invention, 4 primer combinations selected from 144 pairs of primer combinations can produce 4 male specific AFLP markers; the AFLP markers are cloned and sequenced to screen out a sequence characterized amplified regions (SCAR) marker molecular marker; and thus, the genetic sex identification method for striped beakfish is established. In the invention, the AFLP method is used for the first time to screen out the sex specific molecular marker for the striped beakfish and convert the sex specific molecular marker for the striped beakfish into the SCAR marker so as to establish the genetic sex identification method for the striped beakfish. The striped beakfish genetic sex identification method has the advantages of accuracy, sensitivity, reliability and the like, has high scientific value for the study onfish sex and has a great significance and high application value for the sex control and selective breeding of fish.

The invention belongs to a preparation field of a cole molecular marker, in particular to a molecular marker linked to recessive genic sterile gene in brassica napus and breeding of a temporary maintainer line homologous with a genic sterile line. The molecular maker is characterized by comprising the following steps of: establishing near-isogenic lines 7-7365AB, 7-736512AB and 7-7365AC of three genes BnMs3, BnMs4 and BnRf by using two types of male sterile lines 7-7365AB in the brassica napus, brassica napus 7-749 and temporary maintainer line materials 7-7365C as raw materials, and then sieving SSR (Simple Tandem Repeat) primers and AFLP (Amplified Fragment Length Polymorphism) primers and transforming a SCAR marker to obtain a codominant SSR molecular marker SR12384 linked to the BnMs3, dominant SCAR markers SM129, SM98, SM113 and codominant SCAR markers SM23 and H3 which are linked to the BnMs4, and a codominant SSR marker D1 linked to the BnRf. The invention can be used for auxiliary selection and breeding of recessive genic sterile markers and can also be used for fine mapping and cloning of sterile genes.

The invention belongs to the field of genetic breeding, and provides a molecular marker of single fruit weight major quantitative trait loci (QTL) of August red pyrus L. fruits and application of the molecular marker. The genetic linkage map of August red is constructed by using SRAP (sequence-related amplified polymorphism), SSR (simple sequence repeats) and AFLP (amplified fragment length polymorphism), phenotype identification on single fruit weight of pyrus L. is combined, and an interval mapping method is used to analyze linkage groups, thus major QTL is detected on the seventh linkage group of August red and can explain 30.9% of characteristics of single fruit weight. The point which is the nearest to the major QTL site Pfw-1 is marked to be EACAMCAC-2000m, and the nearest distance is 1.019cM. The obtained titratable acid major QTL molecular marker has significant theoretical and practical means for quickening genetic improvement process of pyrus L. varieties and improving breeding selection efficiency.

The present invention relates to a high throughput method for the identification and detection of molecular markers wherein restriction fragments are generated and suitable adaptors comprising (sample-specific) identifiers are ligated. The adapter- ligated restriction fragments may be selectively amplified with adaptor compatible primers carrying selective nucleotides at their 3' end. The amplified adapter-ligated restriction fragments are, at least partly, sequenced using high throughput sequencing methods and the sequence parts of the restriction fragments together with the sample-specific identifiers serve as molecular markers.

The invention discloses an anther culture method for indica japonica hybrid rice. The method comprises the following steps: sampling, identifying a microspore development period, low-temperature pretreatment, young wheatear sterilization, anther inoculation, still dark cultivation at 25+ / - 2 DEG C until the diameter of embryonic callus is 2-3mm; carrying out a redifferentiation stage to form complete regenerated plants with haploid or double haploids. The double haploids are good breeding materials, and can be directly used for breeding of new varieties or varietal improvement; the breeding age limit is greatly shortened; the selection efficiency is improved; the double haploids also are ideal materials for molecular markers, such as amplified fragment length polymorphism (AFLP), restricted fragment length polymorphisms (RFLP), randomly amplified polymorphic deoxyribonucleic acid (RAPD) and the like, genetic map drawing, and researches on animal evolution, genetic analysis, plant gene cloning and screening, genetic engineering breeding and the like.

The invention provides a primer pair consisting of nucleotide sequences shown as SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.3. The primer pair can be applied to the detection of a rose cockscomb character. The invention also provides a method for detecting the rose cockscomb character by using the primer pair. By the method, an obvious correlation property of amplified fragment length polymorphism and the rose cockscomb character in an intron 3 of a coiled-coil domain containing 108 (CCDC108) gene of a chicken is discovered and an amplified fragment can be used as a genetic marker of the chicken,so a molecular biological method is used for auxiliary selection of the marker.

The invention relates to a plant restoration technology, and particularly discloses a method for screening manganese / cadmium absorption efficient ecological phytolacca americana. The method comprises the following steps: collecting in the wild leaves and seeds of phytolacca americana populations in a plurality of contaminated regions and uncontaminated regions in China and soil in the vicinity; detecting the manganese / cadmium content of the soil in the vicinity of the phytolacca americana populations in the regions, and selecting phytolacca americana populations in highly manganese contaminated, highly cadmium contaminated and uncontaminated regions respectively; performing AFLP (amplified fragment length polymorphism) molecular marker detection on the leaves of the selected phytolacca americana populations, and analyzing genetic structures thereof; selecting bred seedlings of the phytolacca americana populations to which different genetic differentiation occurs, culturing the seedlings for 50 days with manganese, cadmium and normal Hoagland solutions, and screening the manganese / cadmium absorption efficient ecological phytolacca americana by adopting the plant heights, fresh weights and leaf manganese / cadmium content of the phytolacca americana as indexes. According to the method, a manganese / cadmium absorption efficient ecological hyperaccumulator is used for treating manganese / cadmium-contaminated soil, and the heavy metal accumulation and restoration efficiency of the hyperaccumulator are improved.

The invention relates to a method for analyzing the genetic diversity of shellfish by using a fAFLP (Fluorescent Amplified Fragment Length Polymorphism) labeling technology, which sequentially comprises the following steps of: extracting DNA of shellfish, digesting the DNA by restriction enzyme, carrying out joint connection on the DNA subjected to the enzyme cutting, carrying out PCR ((Polymerase Chain Reaction) pre-amplification and selective amplification, detecting a product and acquiring and analyzing an image and data. Compared with the prior art, the invention has the advantage of being capable of efficiently, stably and accurately analyzing the genetic diversity of the shellfish by adopting a self-creating reaction system combining with the fAFLP labeling technology. In addition, the invention has the advantages of more amplification sites, high polymorphic site proportion and large Nei genetic diversity.

The invention relates to a method for establishing and optimizing an exopalaemon modestus SRAP(sequence-related amplified polymorphism)-PCR(polymerase chain reaction) system, belonging to the technical field of fisheries biology. The method for establishing and optimizing an exopalaemon modestus SRAP-PCR system is characterized in that 25mu L of the PCR system contains 2mmol / L of Mg<2+>, 0.5U of Taq enzyme, 75ng of template DNA (deoxyribonucleic acid), 0.20mmol / L of dNTPs and 0.2mu mol / L of primers, wherein the primers are 5'-TGAGTCCAAACCGGAGC-3' and 5'-GACTGCGTACGAATTTGA-3'. The currently reported molecular markers applied to exopalaemon modestus blastogensis analysis are only RAPD (randomly amplified polymorphic DNA) and AFLP (amplified fragment length polymorphism). The invention develops a new molecular marker (SRAP molecular marker), and supplements and enriches the means and methods for molecular biological researches on exopalaemon modestus.

The invention discloses a method for obtaining genus hybrid of forsythia and abeliophyllum. The method comprises the following specific steps: (1) carrying out generic artificial hybridization by taking fructus forsythiae of the forsythia as a female parent and taking abeliophyllum distichum of the abeliophyllum as a male parent, and gathering the seeds obtained after the hybridization; and (2) planting the seeds to obtain filial generation materials, carrying out morphological observation statistics by referring to the male parent and the female parent, carrying out hybrid identification by using an AFLP (Amplified Fragment Length Polymorphism) molecular marker and selecting the genus hybrid of the forsythia and the abeliophyllum. According to the method, distant hybridization is carried out on fructus forsythiae of the forsythia and abeliophyllum distichum of the abeliophyllum, so that the generic gene exchange between the forsythia and the abeliophyllum is realized and a generic hybridization new germplasm is obtained. Based on the method, the genetic information of the forsythia plants can be enriched, the new variety cultivation of fructus forsythiae via generic hybridization is possible and the research of affinis genus genetic relationship can be also carried out.

The invention relates to a molecular marker for the content of soluble sugar in an 'August red' pear fruit, which belongs to the field of inheritance breeding. A genetic linkage map of the 'August red' pear fruit is constructed by using SRAP (Sequence-Related Amplified Polymorphism), SSR (Simple Sequence Repeats) and AFLP (Amplified Fragment Length Polymorphism), a colony is analyzed by using an interval mapping method in combination with phenotypic identification of the content of the soluble sugar in the fruit, and the existence of major QTL (Quantitative Trait Loci) on a seventh linkage group of the 'August red' pear fruit is detected, so that the characteristic of the soluble sugar content of 12.4 percent can be explained. A marker which is closest to the locus Psg-1 of the major QTL is me6pm19-1300, wherein the distance is 0 centimeter. The obtained major QTL molecular marker of the soluble sugar plays an important theoretical and practical guide role in accelerating the genetic improvement process of a pear variety and improving the breeding selection efficiency.

The invention relates to a molecular marker seed-breeding technology of corn and solanaceous vegetables. The technology is mainly characterized by implementing researches of technologies, such as extraction and purification of DNA of crops, PCR (Polymerase Chain Reaction) and characteristic primer screening, RAPD (Random Amplified Polymorphic DNA) marking, SSR (sympathetic skin response) marking, AFLP (Amplified Fragment Length Polymorphism) marking, molecular marker-assisted selection of disease-resistance breeding materials, separation and DNA sequencing of target genes and the like aiming at crops, such as corn, tomato, eggplant, chili and the like.

The invention discloses a method for identifying different brewing barley genetic resources at home and abroad by combining a DNA (Deoxyribose Nucleic Acid)-AFLP (Amplified Fragment Length Polymorphism) molecular marker with biological analyzing software. The method is characterized by comprising the following steps of: establishing a plant genome DNA extracting method suitable for AFLP and a barely germ plasma analyzing AFLP system, and carrying out electrophoresis and silver staining onto an amplified product on 6% of denatured polyacrylamide gel; establishing an AFLP fingerprint spectrum of 27 brewing barely varieties at home and abroad by utilizing three pairs of EcoRI / MseI primer combinations (E8 / M1, E8 / M3, E8 / M5) AFLP. By carrying out statistic analysis through banding pattern analyzing software POPGEN, NTSYS 3.0 and STATISTICA / w5.0, the unique banding patterns of 27 materials can be mutually distinguished from other varieties by utilizing 10 specific sites according to electrophoretogram, so that foundation is laid up for furthermore malt production and beer process.

The invention discloses a PCR-RFLP (Polymerase Chain Reaction-Amplified Fragment Length Polymorphism) assay kit for drug resistance of anopheles sinensis and a special primer of the same. The invention provides the special primer which is used for detecting a mutational site of a kdr gene in the anopheles sinensis. The special primer consists of a primer set A and a primer set B, wherein the primer set A consists of a primer 1 and a primer 2, nucleotide sequence of the primer 1 is a sequence 1in a sequence table, and that of the primer 2 is a sequence 2 in the sequence table; and the primer set B consists of a primer 3 and a primer 4, the nucleotide sequence of the primer 3 is a sequence 3 in the sequence table, and that of the primer 4 is a sequence 4 in the sequence table. Based on experiments, the invention provides a quick, simple, convenient and flexible assay kit for kdr mutant allelic genes of the anopheles sinensis, which is used for mastering conditions of kdr allelic genes of the anopheles sinensis relative to the resistance of a pyrethroid pesticide.

The invention discloses a method for identifying Qianshan No.2, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz. by utilizing AFLP (amplified fragment length polymorphism)-CE (capillary electrophoresis) technique, belonging to the technical field of molecular markers. The method comprises the steps of extraction of sample DNA, double digestion of DNA samples and linking of double digestion products, pre-amplification of linking products, selective amplification, AFLP-CE detection and result analysis and identification. By adopting the method, an AFLP-CE separation diagram of Qianshan No.2, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz. is successfully established, whether the samples are Qianshan No.2, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz. can be easily and accurately identified and scientific bases are provided for protecting Qianshan No.2, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz..

The invention provides an erigeron breviscapus DNA polymorphism detection method and application thereof. In the invention, an amplified fragment length polymorphism technology is used for erigeron breviscapus DNA polymorphism detection, an optimum primer combination is screened out, and reaction condition is optimized. The method can be effectively applied to the inheritance breeding, the resource protection, the introduction domestication, the breed optimization and the like of erigeron breviscapus.

The present invention relates to the technical field of molecular biology, provides a method for measuring short RNA using amplified DNA fragment length polymorphism, and comprises the following steps: first using at least two synthesized miRNAs as the internal measurement standard, said synthesized miRNAs containing no natural homologous sequence in comparison with the short RNA to be measured, and mixing the synthesized miRNAs using different molecule numbers so as to form a dynamic miRNA standard molecular gradient; mixing the same quantity of the dynamic miRNA standard with the short RNA to be measured, and performing RNA reverse transcription, cDNA tailing, PCR synchronous amplification, and fluorescent quantitative analysis on the length polymorphism fragment of the PCR product DNA so as to measure the relative ratio of the fluorescence intensity of the DNA fragment produced by the amplification of the short RNA to be measured to the dynamic miRNA standard fluorescence intensity gradient.

The invention discloses a method for identifying Qianshan No.1, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz. by utilizing AFLP (amplified fragment length polymorphism)-CE (capillary electrophoresis) technique, belonging to the technical field of molecular markers. The method comprises the steps of extraction of sample DNA, double digestion of DNA samples and linking of double digestion products, pre-amplification of linking products, selective amplification, AFLP-CE detection and result analysis and identification. By adopting the method, an AFLP-CE separation diagram of Qianshan No.1, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz. is successfully established, whether the samples are Qianshan No.1, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz. can be easily and accurately identified and scientific bases are provided for protecting Qianshan No.1, a new variety of Erigeron Breviscapus (Vant.)Hand.-Mazz..

The invention discloses an establishment method and a use of wild bluegrass AFLP (Amplified Fragment Length Polymorphism) fingerprint. The establishment method comprises six steps of genome DNA (Deoxyribose Nucleic Acid) extraction, enzyme digestion, connection, pre-amplification, selective amplification and denaturing polyacrylamide gel electrophoresis, and then is capable of obtaining a large number of high and low different DNA strips; bluegrass germplasms having the DNA strips of same size are considered to have the same genetic loci and bluegrass germplasms having the DNA strips of different sizes are considered to have different genetic loci, and on this basis, the fingerprint of different wild bluegrass germplasms is established, and can be used for identifying the bluegrass germplasms, identifying variety and protecting the germplasms. The establishment method has the advantages that the genome DNAs of the bluegrass plants are directly detected and the influence on the phenotypic characters of the genome DNAs caused by factors such as the field environment is avoided, and in addition, the method can be used for detecting and identifying the genome DNAs at the seedling stage, and therefore, the detection period is shortened.

Particular aspects provide an inexpensive QTL mapping approach for genome-wide scans of QTL linked markers and for narrowed locations of QTL regions, which may comprise integration of the amplified fragment length polymorphism (AFLP) with DNA pooling and selective genotyping and comparative bioinformatics tools. AFLP simultaneously screens high numbers of loci for polymorphisms and detects many more polymorphic DNA markers than any other PCR based detection systems, and provides for identification of “responsible mutations” for a QTL. Aspects of the present invention also provide novel compositions and methods based on novel DOPEY2 and / or KIAA1462 single nucleotide polymorphisms such as AAFC03071397.1:g.12881G>C, g.12925G>A, g.12951T>C, g.13013A>G, g.13125G>A and g.13173C>T in the DOPEY2 gene and AAFC02113318.1:g.1367G>A and g.1372G>A in the KIAA1462 gene, which may provide novel markers for beef marbling and subcutaneous fat. Additional aspects provide for novel methods which may comprise marker-assisted selection to improve beef marbling and subcutaneous fat in cattle.

The invention relates to a primer set for identifying a Chinese prawn population. The primer set comprises four pairs of primers, wherein sequences of a forward primer and a reverse primer of each pair of primers are SEQ ID NO: 1 and SEQ ID NO: 2, SEQ ID NO: 1 and SEQ ID NO: 3, SEQ ID NO: 4 and SEQ ID NO: 5, and SEQ ID NO: 6 and SEQ ID NO: 7 respectively. According to a primer combination screened by the invention, the source and the place of the Chinese prawn population at a natural sea area can be determined, and the contribution of different spawning sites to the Chinese prawn population is favorably defined. Chinese prawns are widely distributed in the sea area shared by China, Japan and Korea; according to the primer combination screened by the invention, the contribution of different spawning water areas to the condominium water area resource can be determined, and resource place proportion and deserved fishing quota can be defined; and the method has great significance to safeguard of Chinese maritime right and protection of Chinese fishery resources.

The invention provides a molecular marker for identifying a mitochondrion gene related to CMS (cytoplasmic male sterility) in a green Chinese onion, a preparation method and an application method thereof. The nucleotide sequence of the molecular marker of the mitochondrion gene related to CMS is shown as SEQ No:1 in a sequence table; and the nucleotide sequences of primers for amplifying the molecular marker are shown as SEQ No:2 and SEQ No:3 in the sequence table. According to the invention, the AFLP (amplified fragment length polymorphism) molecular marker of the mitochondrion gene related to CMS in a green Chinese onion is converted into the SCAR (sequence characterized amplified region) molecular marker, so that a large number of samples can be quickly detected, the result is good in stability, the reproducibility is high, and the operation is facilitated. The SCAR marker in the invention can detect all varieties in the experiment, namely 8 sterile line varieties, 8 maintainer line varieties and 4 hybrid varieties; and the marker has a wide application range, can conveniently identify the cytoplasm type of the green Chinese onion and assists the development of molecular breeding.

The present invention relates to the technical field of molecular biology, provides a method for measuring short RNA using amplified DNA fragment length polymorphism, and comprises the following steps: first using at least two synthesized miRNAs as the internal measurement standard, said synthesized miRNAs containing no natural homologous sequence in comparison with the short RNA to be measured, and mixing the synthesized miRNAs using different molecule numbers so as to form a dynamic miRNA standard molecular gradient; mixing the same quantity of the dynamic miRNA standard with the short RNA to be measured, and performing RNA reverse transcription, cDNA tailing, PCR synchronous amplification, and fluorescent quantitative analysis on the length polymorphism fragment of the PCR product DNA so as to measure the relative ratio of the fluorescence intensity of the DNA fragment produced by the amplification of the short RNA to be measured to the dynamic miRNA standard fluorescence intensity gradient.

The invention provides a molecular marker of the Pyrus communis L. fruit titratable acid content, belonging to the genetic breeding field. Sequence related amplified polymorphism (SRAP), simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) are utilized to construct the genetic linkage maps of Pyrus communis L., the phenotypic identification of the fruit titratable acid content is combined, the interval mapping method is adopted to analyze the colony, a main effect quantitative trait locus (QTL) exists in the third linkage group of Pyrus communis L. through detection, and the characteristic that the titratable acid content is 49.3% can be explained. The marker EAACMCAG-850m* is most near the main effect QTL Pta-1, and the distance between the marker and the locus is 2cm. The obtained molecular marker of titratable acid main effect QTL has important theoretical and practical significances for accelerating the genetic improvement process of Pyrus L. cultivars and increasing the breeding selection efficiency.

The invention provides a striped beakfish sex specific molecular marker and a genetic sex identification method thereof. The method comprises: screening and cloning of a male specific amplified fragment length polymorphism (AFLP) marker; performing DNA sequence analysis on sex specificity; and constructing a genetic sex polymerase chain reaction (PCR) identification method. In the invention, 4 primer combinations selected from 144 pairs of primer combinations can produce 4 male specific AFLP markers; the AFLP markers are cloned and sequenced to screen out a sequence characterized amplified regions (SCAR) marker molecular marker; and thus, the genetic sex identification method for striped beakfish is established. In the invention, the AFLP method is used for the first time to screen out the sex specific molecular marker for the striped beakfish and convert the sex specific molecular marker for the striped beakfish into the SCAR marker so as to establish the genetic sex identification method for the striped beakfish. The striped beakfish genetic sex identification method has the advantages of accuracy, sensitivity, reliability and the like, has high scientific value for the study onfish sex and has a great significance and high application value for the sex control and selective breeding of fish.

The present invention belongs to the field of breeding new plant variety, and features that through the breeding process including one cross breeding, three backcrosses and one selfing, the destination gene is introduced into hybrid rice restoring line to be improved. By means of the detection of polymerase chain reaction and the selection of molecular marker gene co separated from and / or closely interlocked with the destination gene, the transferring fragment containing the destination gene is made to be as small as possible. By applying augmented fragment length polymorphism label in selecting genetic background, the improved restoring line has the same genome the same as original restoring line except for the destination gene fragment.

The invention provides a molecular marker of the Pyrus communis L. fruit titratable acid content, belonging to the genetic breeding field. Sequence related amplified polymorphism (SRAP), simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) are utilized to construct the genetic linkage maps of Pyrus communis L., the phenotypic identification of the fruit titratable acid content is combined, the interval mapping method is adopted to analyze the colony, a main effect quantitative trait locus (QTL) exists in the third linkage group of Pyrus communis L. through detection, and the characteristic that the titratable acid content is 49.3% can be explained. The marker EAACMCAG-850m* is most near the main effect QTL Pta-1, and the distance between the marker and the locus is 2cm. The obtained molecular marker of titratable acid main effect QTL has important theoretical and practical significances for accelerating the genetic improvement process of Pyrus L. cultivars and increasing the breeding selection efficiency.