Fluorescence in situ hybridization hTERT transfected external quality assessment cell line and preparation method thereof

Abstract

The invention relates to a fluorescence in situ hybridization hTERT transfected external quality assessment cell line for diagnosis quality control in medical genetics, and a preparation method thereof. The invention is mainly characterized in that the preparation method comprises: performing double digestion of plasmid pCIneo-hTERT and a carrier pLXSNneo by endonucleases of EcoR I and Xho I, connecting the digestion products of hTERT and pLXSNneo by Ligation Mix, constructing a pLXSNneo-hTERT recon, transfecting, by liposome, residual adherent living cells which are in logarithmic growth and are definitely diagnosed to have common numerical abnormalities of chromosomes as needed by clinical diagnosis and treatment, screening cell lines integrated with the recons by G418, performing subculture amplification and cryopreservation, then taking the cell lines, mixing the cell lines according to a required ratio for quality control so as to convert each original cell line with only one numerical abnormality of chromosomes into a chimera quality-control cell line with a certain ratio of common numerical abnormalities of chromosomes. Therefore, the difficulty for differential diagnosis and chimera diagnosis is increased; original cells are easily available; waste residual cells are converted into effective immortalized quality-control cells; external quality assessment is carried out by the difficult quality-control cells; and the cell line of the invention has important significance on in-time discovery and solution of quality problems, and diagnostic level improvement.

Description

technical field

[0001] The invention relates to a fluorescent in situ hybridization hTERT transfected cell line for inter-laboratory quality assessment and a preparation method thereof, which is mainly used in molecular genetics diagnostic laboratories in the medical field for technical assessment of fluorescent in situ hybridization diagnosis, indoor quality control and inter-laboratory quality control. Quality review. Background technique

[0002] Fluorescent in situ hybridization is mainly used to detect abnormalities in the number and structure of chromosomes. Chromosomes are the genetic material in the nucleus. Humans have 23 pairs of chromosomes, 22 of which are autosomes, and 1 pair is the sex chromosome that determines the sex of men and women. Genes carrying genetic information on chromosomes, of which DNA accounts for more than 90%, and RNA content varies with cell cycle and growth, generally accounting for 1% to 10%.

[0003] Chromosomal structural abnormalities,...

Examples

Embodiment Construction

[0018] 1. Primordial cells: The primitive cells used are derived from trisomy 21, trisomy 18, trisomy 13, X or Y chromosome after diagnosis of chromosomal disease due to the needs of clinical diagnosis and treatment and have been diagnosed as one of the common abnormalities in chromosome number Abnormal remaining primary or passage living tissue cells that grow adherently, and the primary or passage living tissue cells may also involve living tissue cells with abnormal chromosome structure or No. 1 to No. Live tissue cells with abnormal number of chromosome 22. The cell types are fetal amniotic fluid, villous tissue and other cells.

[0019]2. Extraction of hTERT: ① Digestion of pClneo-hTERT: hTERT is located between the EcoRI and SalI sites of the plasmid pClneo-hTERT, and the multiple cloning site (MCS) of the pLXSNneo vector contains EcoRI and XhoI restriction sites. Take the pCIneo-hTERT plasmid and dissolve it in an appropriate amount of ultra-clean H 2 In O or TE buffe...

technical field

[0001] The invention relates to a fluorescent in situ hybridization hTERT transfected cell line for inter-laboratory quality assessment and a preparation method thereof, which is mainly used in molecular genetics diagnostic laboratories in the medical field for technical assessment of fluorescent in situ hybridization diagnosis, indoor quality control and inter-laboratory quality control. Quality review. Background technique

[0002] Fluorescent in situ hybridization is mainly used to detect abnormalities in the number and structure of chromosomes. Chromosomes are the genetic material in the nucleus. Humans have 23 pairs of chromosomes, 22 of which are autosomes, and 1 pair is the sex chromosome that determines the sex of men and women. Genes carrying genetic information on chromosomes, of which DNA accounts for more than 90%, and RNA content varies with cell cycle and growth, generally accounting for 1% to 10%.

[0003] Chromosomal structural abnormalities,...

Examples

Embodiment Construction

[0018] 1. Primordial cells: The primitive cells used are derived from trisomy 21, trisomy 18, trisomy 13, X or Y chromosome after diagnosis of chromosomal disease due to the needs of clinical diagnosis and treatment and have been diagnosed as one of the common abnormalities in chromosome number Abnormal remaining primary or passage living tissue cells that grow adherently, and the primary or passage living tissue cells may also involve living tissue cells with abnormal chromosome structure or No. 1 to No. Live tissue cells with abnormal number of chromosome 22. The cell types are fetal amniotic fluid, villous tissue and other cells.

[0019]2. Extraction of hTERT: ① Digestion of pClneo-hTERT: hTERT is located between the EcoRI and SalI sites of the plasmid pClneo-hTERT, and the multiple cloning site (MCS) of the pLXSNneo vector contains EcoRI and XhoI restriction sites. Take the pCIneo-hTERT plasmid and dissolve it in an appropriate amount of ultra-clean H 2 In O or TE buffe...

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