32 results about "Cytoplast" patented technology

This invention discloses a rapid method for creating wheat CMS maintainer lines based on recurrent selection and double haploid technology. The method involves crossing a CMS sterile line with a fertile cytoplasmic restorer line to obtain heterozygous fertile plants. Then, using the restorer line as the recurrent paternal parent, a backcross is performed to convert the cytoplasm to a fertile genotype. Double haploid technology is used to rapidly obtain a homozygous DH population from the backcross progeny. Finally, through testcross identification, DH plants with completely sterile cytoplasmic cytoplasm are selected, thus obtaining the target maintainer line with fertile cytoplasm and a sterile nuclear genotype. This invention further incorporates recurrent selection technology for continuous optimization of the maintainer line's agronomic traits. This invention integrates backcrossing, DH technology, and a recurrent selection system, significantly shortening the breeding cycle. The maintainer line exhibits homozygosity ≥99%, a pure cytoplasmic background, strong versatility, and applicability to wheat CMS types, providing efficient technical support for hybrid wheat breeding.

This invention discloses a specific molecular marker for identifying eggplant Saet CMS cytoplasmic male sterility lines and its application. The molecular marker, Saet218b, was selected by comparing the mitochondrial genomes of the eggplant Saet CMS cytoplasmic male sterility line and its maintainer line. Through bioinformatics analysis, the candidate specific gene for Saet CMS male sterility, orf218b, was obtained, and its nucleotide sequence is shown in SEQ ID NO: 1. Based on this sequence information, the molecular marker Saet218b for identifying eggplant Saet CMS cytoplasmic male sterility lines was successfully designed. The molecular marker of this invention can be used to identify eggplant Saet CMS cytoplasmic male sterility lines efficiently and with a simple procedure. The results can be used for marker-assisted breeding of eggplant Saet-derived cytoplasmic male sterility lines, improving breeding efficiency. It also provides technical support and theoretical basis for the identification of eggplant specific cytoplasmic germplasm.

The invention relates to a lung fibroblast culture method, and belongs to the technical field of cell culture. Comprising the following steps: S1, organization preparation; s2, tissue treatment; s3, primary culture: the treated tissue blocks are attached to a culture dish, a culture solution is added, the culture solution is a primary culture system, and culture is performed in an incubator for several days until the culture dish is full of fibroblasts; s4, subculture: digesting the cells subjected to primary culture by adopting conventional 0.25% trypsin, then adding a culture solution which is a subculture system for subculture, and carrying out subculture to a fourth generation; and S5, cell identification. According to the invention, primary and subculture are respectively purified and amplified by virtue of a double culture system of fibroblasts, so that the purification of target cells is facilitated, the cell quality is good, a low-serum growth additive is used as a nutrient substance, the low-serum growth additive is more easily utilized by the cells, the proliferation of the cells is facilitated, and good proliferation activity and phenotypic stability are still kept when the cells are subcultured to the fourth generation; no obvious differentiation phenomenon occurs.

This invention belongs to the field of plant genetic engineering, specifically disclosing a method for creating bifunctional haploid inducible lines of Brassica napus based on the BnaKNL2 gene and its application. Research revealed that the BnaKNL2 gene is a key gene affecting the fertility of Brassica napus. By knocking out the KNL2 gene in Brassica napus, bifunctional haploid inducible lines with maternal or paternal haploid induction capabilities can be created. The haploid induction rate after hybridization with different Brassica napus varieties can reach 0.45%–2.7%. Furthermore, by combining this with a cytoplasmic male sterility system and through hybridization and backcrossing, the CMS-HI line, carrying both sterile cytoplasm and haploid induction capabilities, can be rapidly created. This provides a new approach for haploid breeding of Brassica napus and the improvement and creation of CMS lines, with broad application prospects.

The invention relates to the cross technical field of molecular biology and parasitology, and particularly discloses a giardia living body marking method based on fluorescent protein UnaG, which comprises the following steps: firstly, optimizing and synthesizing a UnaG gene sequence according to the codon use preference of giardia; then, constructing a marker plasmid pGT-UnaG-neo on a double expression framework containing an alpha2-tubulin promoter and a gdh promoter, and introducing the plasmid into the giardia giardia trophozoite through electroporation. And carrying out resistance screening in a culture medium containing geneticin to obtain an insect strain capable of stably integrating and expressing UnaG. Through bilirubin induction, efficient maturation and green fluorescence emission of the UnaG protein in giardia cytoplasm are realized. The stably transferred strain obtained by the method has no significant difference from a wild strain in the aspects of growth curve and survival rate, and UnaG and bilirubin have no obvious toxic effect. The limitation that traditional fluorescent protein is inactivated under the anaerobic condition is broken through, and a reliable tool is provided for dynamic imaging, functional research and drug screening of giardia and other anaerobic protozoa.

A cell quality evaluation apparatus performs a process including estimation processing of estimating a quality of cells in the entirety of a cell-culture container by determining feature quantities from a plurality of images and calculating an average value of the feature quantities; derivation processing of deriving an estimation error of the quality estimated in the estimation processing, based on a variation of the feature quantities in the plurality of images and imaging information related to an area of a plurality of imaging regions; and imaging control processing of causing a imaging apparatus to perform re-imaging on at least one re-imaging region different from the plurality of imaging regions in a case where the estimation error is out of an allowable range.

This invention relates to the field of biomedical technology and discloses a biomarker for diagnosing polycystic ovary syndrome (PCOS) and its application. The diagnostic biomarker is circSPECC1(4), which is formed by reverse splicing and circularization of the fourth exon of the SPECCC1 gene. It has a length of 1580 nt and possesses a closed circular RNA structure. circSPECC1(4) is specifically highly expressed in ovarian granulosa cells of PCOS patients, and there is no significant difference in the mRNA expression level of its parent gene SPECCC1. circSPECC1(4) possesses RNase R nuclease resistance stability and is mainly located in the cytoplasm. Knocking down the expression of circSPECC1(4) can significantly promote apoptosis of ovarian granulosa cells and arrest the ovarian granulosa cell cycle at the G0 / G1 phase. This invention clearly demonstrates that circSPECC1(4) can serve as a specific diagnostic biomarker for PCOS, fully verifying the structural characteristics, expression specificity, and pathological regulatory function of this circular RNA, filling the gap in the existing field of PCOS diagnosis which lacks highly specific and stable molecular diagnostic targets.

This invention belongs to the field of plant molecular breeding, and specifically discloses a gene related to rice fertility restoration. OsRf19 This invention addresses the cumbersome process of identifying restorer lines for the existing Fujian abortive cytoplasmic male sterility (CMS-FA / OsRf19) system by developing a method for creating tightly linked artificial haplotypes, the development of InDel molecular markers, and their application in rice fertility. Through bioinformatics analysis, this invention identifies a fertility restoration gene associated with the rice sporophytic sterility system CMS-FA. OsRf19 A specific region closely linked to the PPR gene cluster and with extremely low similarity to the loci was edited using CRISPR / Cas12a gene editing technology, resulting in multiple large deletions. Based on these genotype sequence variations, a set of InDel molecular markers was developed to realize the editing of this artificially created restoration gene. OsRf19 Rapid identification of closely linked artificial haplotypes accelerates the breeding process of molecular marker-assisted selection in the rice CMS-FA system.

The invention belongs to the field of gene engineering and molecular breeding, and particularly discloses interaction protein MORF8 of rice wild abortion type cytoplasmic male sterility recovery protein Rf4 and application of the interaction protein MORF8, the amino acid sequence of the interaction protein MORF8 is shown as SEQ ID NO.2, the interaction protein MORF8 is coded by an MORF8 gene, and the nucleotide sequence of the MORF8 gene is shown as SEQ ID NO.1; the application of the interaction protein MORF8 of the rice wild abortion type cytoplasmic male sterility restoring protein Rf4 in rice breeding comprises the following steps: constructing a rice wild abortion type cytoplasmic male sterility line overexpressing the MORF8, and hybridizing the rice wild abortion type cytoplasmic male sterility line overexpressing the MORF8 with a rice wild abortion type cytoplasmic male sterility restoring line containing the restoring protein Rf4, hybrid seeds which synergistically enhance the fertility recovery capability of the recovery protein Rf4 are generated. Compared with the prior art, the invention discloses a synergistic enhancement function of MORF8, obviously improves the maturing rate of rice, and provides an efficient and safe novel molecular breeding tool for creating high-yield and stable-yield hybrid rice.

This invention provides molecular markers, typing primers and their applications for the restoration of cytoplasmic male sterility in chili pepper. Wild-type and mutant chili pepper materials were used to construct an F2 population, and the chromosomal region closely linked to the cytoplasmic male sterility restoration gene in chili pepper was obtained using the BSA population positioning method. A KASP molecular marker was designed based on a single base mutation, and this marker was used to identify the genotypes of 1290 plants in the F2 population, with a match rate of 100%. This invention is not only useful for the selection and assisted breeding of chili pepper for cytoplasmic male sterility, but also provides a basis for the mapping of the cytoplasmic male sterility restoration gene and the analysis of the molecular mechanism of cytoplasmic male sterility, and has widespread value.

Downregulated miR-99b-5p and upregulated mTOR cooperatively promotes the African American (AA) PCa aggressiveness and drug resistance. Nuclear mTOR, AR, and SMARCD1 are highly expressed in AA PCa (MDA PCa 2b) compared to EA PCa (LNCaP) cell line. miR-99b-5p inhibited protein levels of mTOR, AR / AR-V7 and SMARCD1 in cytoplasm and nuclei of EA and AA PCa. miR-99b-5p effectively inhibits cell proliferation / survival and induced cell apoptosis in EA and AA PCa cells. Moreover, combination of miR-99b-5p and enzalutamide (Enz) synergistically enhances the cytotoxicity against aggressive AA PCa and castration resistant prostate cancer (CRPC). miR-99b-5p or miR-99b-5p / Enz significantly reduces the recruitment of mTOR to the genes involved in the metabolic reprogramming in CRPC. miR-99b-5p can function as an epigenomic driver to modulate the mTOR / AR / SMARCD1 signaling axis in AA PCa and resistant CRPC. miR-99b-5p can be utilized as a biomarker for identifying the presence of prostate cancer.

The application relates to a stem cell culture density optimization control method and system, and relates to the technical field of stem cell culture, and comprises the following steps: K culture boxes and P culture dishes are configured; when mesenchymal stem cells are cultured, the P culture dishes are randomly and equally distributed to the K culture boxes; P cell imaging sequences in a historical time window are monitored; P real-time cell proliferation rates and morphological characteristics are obtained through analysis; the P culture dishes are similar clustered based on the same; K similar culture dish sets and K cell proliferation rate averages are determined; the K similar culture dish sets are mapped and placed in the K culture boxes; and the K culture boxes are respectively optimized and adjusted in culture environment parameters based on the K cell proliferation rate averages, so that K adaptive environment parameters are obtained, and the K culture boxes in a future time window are controlled. The application solves the problems that in traditional stem cell culture, a unified culture environment is adopted, targeted culture is lacked, and the culture density control precision is low and the cell quality is unstable.

The application belongs to the technical field of genetic engineering, and particularly relates to a negative-sense RNA virus plant genome editing system and application thereof. The genome editing system uses a CRISPR-Cas9 gene editing technology, takes a plant cytoplasmic rhabdovirus as a delivery carrier, is transmitted to a plant by means of a vector, utilizes an RNA movement motif, delivers gene editing components to an axillary meristem, and obtains mutant lateral buds along with plant tillering. The application can complete plant gene editing without tissue culture, overcomes plant tissue culture dependence, expands the range of gene editing lines, and only needs one generation to obtain a pure mutant without going through hybridization and selfing processes.

The present disclosure provides a cytoplasmic male sterile Rudbeckia plant. The present disclosure also provides a method for conferring cytoplasmic male sterility to a Rudbeckia plan. The method may comprise introducing, as a cytoplasmic male sterility gene, at least one polynucleotide into a Rudbeckia plant of interest.

The application discloses a method for breeding and multiplying a flue-cured tobacco cytoplasmic male sterile line MS Guiyan 19, which takes the flue-cured tobacco cytoplasmic male sterile line MSG28 as a cytoplasmic male sterile gene source, takes MSG28 as a female parent, takes a variety Guiyan 19 as a recurrent parent, and is bred into a new cytoplasmic male sterile line MS Guiyan 19 through continuous backcrossing for six generations after hybridization; and the cytoplasmic male sterile line MS Guiyan 19 is bred by taking the sterile line MS Guiyan 19 as the female parent and taking the maintainer line Guiyan 19 as the male parent and is bred and preserved through hybridization. The flue-cured tobacco sterile line MS Guiyan 19 bred by the application has the advantages of stable sterility, excellent agronomic characters, higher yield and output value, good easy-curing and curing properties, good appearance and quality of tobacco leaves, and coordinated chemical components, and provides a new excellent sterile line germplasm for flue-cured tobacco hybridization advantage utilization research work.

Described herein is the discovery that neither the nuclear localization signal (NLS) nor the prion-like domain (PLD) of TDP-43 is necessary for embryonic stem cell culture and differentiation into motor neurons in vitro. The ability of ES cells to express these TDP-43 mutants and differentiate into motor neurons that exhibit an ALS-like phenotype whereby the TDP-43 mutants redistribute to and aggregate in the cytoplasm and fail to regulate cryptic exon splicing allows these cells to act as a model of TDP-43 proteinopathy for the testing of candidate therapeutic agents that may resolve such proteinopathy. Additionally, these ES cells may be used to successfully generate non-human animals, e.g., mice, that also exhibit hallmark symptoms of ALS and that may be used in testing candidate agents useful in treating TDP-43 proteinopathies.

In some aspects, methods are provided for generating primordial follicles from human and other mammalian germ cells with high efficiency, high cell quality, and high reproducibility. In some aspects, methods and culture systems are provided for generating an ovarian organoid, such as a cell aggregate consisting of both somatic and germ cells from a source derived from primary and / or stem cells, the ovarian organoid is capable of progressing primordial germ cells into oocytes that initiate meiosis and are capable of follicular assembly. In some aspects, provided embodiments include the step of culturing an ovarian organoid in a fully defined serum-free medium and under conditions that generally employ a low defined serum protein component.

Providing a new method for determining gene mutations in JMML. [Solution] A method for determining gene mutations in juvenile myelomonocytic leukemia (JMML), comprising the step of determining that the subject has gene mutations in the CBL group or NF1 group if the distribution of monocyte morphology in a blood sample taken from the subject is as follows: (1) if there is a lot of A and a little D, the subject is in the PTPN11_somatic group; (2) if there is a lot of B, the subject is in the PTPN11_germline group; (3) if there is a lot of C, the subject is in the KRAS group; (4) if there is little A and C and a lot of B and D, the subject is in the NRAS group; or (5) if the subject does not fall into any of the above categories (1) to (4), the subject is in the CBL group or NF1 group. A is a monocyte that has cytoplasmic vacuoles and a club-shaped nucleus; B is a monocyte that does not form a cytoplasmic vacuole and has a club-shaped nucleus; C is a monocyte that has cytoplasmic vacuoles and a round nucleus; D represents a monocyte that does not form a cytoplasmic vacuole and has a round nucleus.

This disclosure provides a method and apparatus for identifying enhancer activity in live cells, enabling sensitive detection of enhancer activity within live cells. The method utilizes a nano-electroporation biochip to perform highly focused perforation of the cell membrane and provides the driving force to accelerate the delivery of enhancer probes to the cell nucleus. The enhancer probes are designed using a cyclic amplification strategy, increasing the fluorescence signal by more than 100-fold within one hour. In the embodiments, the method and apparatus successfully detected the activity of CCAT1 enhancer in live cells without affecting cell viability, demonstrating the robustness and reliability of the proposed method and apparatus in detecting enhancer activity in live cells. This disclosure provides a convenient and universal platform for research on enhancer gene regulatory mechanisms in live cells and for clinical diagnostic applications.

Provided herein are cytoplasts, compositions comprising cytoplasts, methods of using cytoplasts, and methods of treating a subject, such as providing benefits to a healthy or unhealthy subject, or treating or diagnosing a disease or condition in a subject. In some embodiments, methods of treating a subject include: administering to the subject a therapeutically effective amount of a composition comprising a cytoplast. Also, provided herein are compositions (e.g., pharmaceutical compositions) that include a cytoplast. Also, provided herein are kits comprising instructions for using the compositions or methods.

This disclosure relates to applications of cell culture, and more specifically to applications of cell culture for growing cells such as stem cells (e.g., PSCs). The method comprises seeding cells in a first container and optionally transferring the cells from the first container to a second container. The cells in the first container, and, if applicable, the cells in the second container, are brought into contact with at least one externally added extracellular matrix or extracellular matrix protein. The method of this disclosure yields a population of cells that are significantly and rapidly grown while maintaining cell quality. The method of this disclosure may also improve the cell seeding efficiency of cells that are significantly and rapidly cultured / grown in a monolayer.

The present application relates to the field of biology, and provides a method for delivering interfering RNA by using cytoplasmic RNA virus as a carrier. The method comprises inserting RNA sequence with RNA cleavage activity at one end or both ends of the interfering RNA sequence by using cytoplasmic RNA virus as a carrier, wherein when the RNA sequence with RNA cleavage activity is inserted at one end of the interfering RNA sequence, a characteristic RNA sequence recognized by RNase is preferably inserted at the other end of the interfering RNA sequence to replace the intranuclear Drosha processing process, increase the cleavage efficiency of the interfering RNA, and achieve the purpose of interfering with the expression of target genes.

The invention provides a method for improving cow superovulation mother cell quality and application thereof, and aims to improve cow superovulation effect and oocyte quality by improving methionine level of oocyte development microenvironment. The method comprises the following steps: feeding rumen protected methionine to the superovulation dairy cow in vivo, and / or adding methionine into a culture solution when in-vitro maturation culture is carried out on oocytes obtained by superovulation. In the embryo level, the blastocyst forming rate and the embryo quality are remarkably improved, the embryonic development potential is higher, and the quality is better. According to the method, by feeding rumen protected methionine in vivo and / or culturing and adding methionine in vitro, the embryo output effect of cow superovulation is systematically improved, the oocyte quality and the embryo development potential are comprehensively improved from the functional and molecular level, and an innovative solution is provided for efficient embryo production.

The invention discloses a steady-state long-term culture method of adipose-derived stem cells. The steady-state long-term culture method comprises the following steps: step 1, obtaining and pretreating adipose tissues; 2, constructing a basic culture system; step 3, regulating and controlling culture in stages; step 4, multi-dimensional steady state monitoring is carried out; 5, precise passage optimization is carried out; 6, whole-process pollution prevention and control; 7, cell quality identification; step 8, low-temperature preservation and recovery; according to the method, EGF, bFGF, TGF-beta1 and a Wnt inhibitor IWR-1 are added in stages, and an antioxidant is combined, so that long-term culture of more than 15 generations of AD-MSC is realized, the positive rate of a dry marker is always maintained to be greater than or equal to 95%, and the differentiation potential is not obviously reduced; a multi-dimensional steady-state monitoring system is established, the components of the culture medium and culture conditions are adjusted in real time, and the problems that the cell quality is not uniform and the inter-batch difference of the cell activity and the proliferation rate is smaller than or equal to 5% due to nutrient fluctuation and metabolic waste accumulation are avoided.

The present application relates to the technical field of single cell RNA sequencing, and discloses a CellRanger single cell RNA sequencing data analysis method, which comprises hierarchical processing of cell quality screening and a tolerant mode of gene expression screening. The method provided by the present application not only strictly guarantees the integrity of various functional indicators in the cells finally stored in the library, but also retains part of the cell information which may have biological significance due to the loss of candidate markers caused by low overall transcriptional activity through further judgment of the low expression state of the cells to be determined. Through this method, the reliability of subsequent analysis data can be ensured, and the information of low expression or rare cells missed due to strict one-size-fits-all standards can be reduced as much as possible.

This application discloses a rapid detection method for the genetic purity of BT-type cytoplasmic male sterile rice seeds based on real-time quantitative PCR. The method primarily utilizes specific primers designed targeting sequences unique to maintainer lines in the rice mitochondrial genome, using the rice internal reference gene ATP6 as a reference to ensure high specificity and accuracy. A standard curve of a mixture of sterile and maintainer line seeds constructed based on this method allows the conversion of ΔCt values ​​into relative abundance for quantitative analysis. Results show that this method exhibits extremely strong linearity, accurate quantification in simulated mixed samples, and a near 100% recovery rate, providing a rapid, stable, and reliable detection method for the quality control of rice hybrids.

An abnormal expression of cytoplasmic Cep63 has been found to be enhanced in tumor cells of which the formation has been promoted due to p62 deficiency. Similarly, the expression of cytoplasmic Cep63 has been found to be enhanced in human pancreatic cancer. On the other hand, suppressing the expression of cytoplasmic Cep63 in the tumor cells and the human pancreatic cancer has been found to inhibit the proliferation thereof. Thus, a new target molecule contributing to p62-related tumor formation has been found, and cancer can be treated or prevented by suppressing the function of the molecule. Cancer can also be detected using the expression of the target molecule as an indicator.

This invention discloses the CsGSTU8 gene of tea plant and its application in alleviating and degrading glyphosate toxicity, relating to the field of genetic engineering technology. The nucleotide sequence of the CsGSTU8 gene is shown in SEQ ID NO.1 of the sequence listing. The amino acid sequence of the protein encoded by the CsGSTU8 gene is shown in SEQ ID NO.2 of the sequence listing. CsGSTU8 is upregulated in both roots and leaves in response to glyphosate stress; CsGSTU8 is specifically highly expressed in tea plant roots. The pCAMBIA1305 plasmid (containing a GFP tag) constructed from this gene, after transformation with Agrobacterium and infection of tobacco leaves, was observed to be localized in the nucleus and cytoplasm. The CsGSTU8 protein is expressed in prokaryotes, and its enzyme activity is strongest at pH 8.0. Transforming the pYES2 plasmid constructed from this gene into the glyphosate-sensitive strain BY4741 restored the growth ability of the yeast mutant on high-concentration glyphosate medium. The cloning of this gene elucidates its role in tea plant resistance to glyphosate stress, providing data support for research on the molecular mechanisms of tea plant resistance to glyphosate stress.

The invention discloses a rice deubiquitination enzyme gene OsZUFSP as well as cloning and application of the rice deubiquitination enzyme gene OsZUFSP. The OsZUFSP is a deubiquitination enzyme gene separated from rice, is composed of 1245 basic groups, and encodes a ZUFSP type deubiquitination enzyme with the molecular weight of 46 kDa and positioned in a cell nucleus and cytoplasm, namely the OsZUFSP. The OsZUFSP participates in the rice ubiquitination removal process and responds to the biological stress response of southern rice black-streaked dwarf virus invasion. The phenotypes of mutant rice with the OsZUFSP gene knocked out and over-expressed transgenic rice are not obviously different from those of wild rice, but the resistance of the mutant rice with the OsZUFSP gene knocked out to the southern rice black-streaked dwarf virus is enhanced, the symptoms of diseased plants are relieved, the susceptibility of the transgenic rice with the OsZUFSP gene over-expressed to the southern rice black-streaked dwarf virus is enhanced, and the resistance of the mutant rice with the OsZUFSP gene knocked out to the southern rice black-streaked dwarf virus is reduced. The symptom of the diseased plant is aggravated, which indicates that the OsZUFSP gene positively regulates and controls the southern rice black-streaked dwarf virus to infect rice Therefore, the gene can be knocked out through a gene editing technology to improve the resistance of the rice to the southern rice black-streaked dwarf virus.

Various methods and compositions are provided for identifying and / or selecting a sorghum plant or germplasm with or without a cytoplasmic male sterility (CMS) trait. In certain embodiments, the method comprises detecting at least one allele of one or more marker locus within or linked to a QTL associated with CMS. In further embodiments, the method comprises crossing a selected sorghum plant with a recurrent sorghum parent plant and selecting progeny with CMS.