CN110283848A - Application of the magnetic induction albumen as magnetic resonance imaging tracer in stem cell tracer - Google Patents
Application of the magnetic induction albumen as magnetic resonance imaging tracer in stem cell tracer Download PDFInfo
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- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
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Abstract
The invention discloses magnetic induction albumen (Magnetoreceptor, MagR) it is used as magnetic resonance imaging (magnetic resonance imaging, MRI) tracer is transduceed into stem cell, observes it in the intracellular overexpression situation of transfection and magnetic resonance imaging effect and the growing state for observing imaging in vivo tracer.In magnetic resonance molecular imaging, magnetic induction protein gene imports stem cell, stem cell is overexpressed and passes through the recruitment to free iron, it is able to the accumulation of iron content in stem cell, realize the gradually amplification of magnetic resonance imaging signal, the sensibility to stem cell signal is improved, to obtain characteristic low signal in magnetic resonance imaging.
Description
Technical field
Application the present invention relates to magnetic induction albumen as magnetic resonance imaging tracer in stem cell tracer, and in particular to
Building magnetic induction prion carrier is simultaneously transduceed into stem cell, observes magnetic induction albumen in the intracellular overexpression situation of transfection and magnetic
Resonance image-forming effect.
Background technique
The development of stem cell regenerating medicine brings a change for modern medicine, and is considered as treatment human tissue, device
Official's damage lacks and the method for the major diseases most prospects such as lesion.Stem cell can be used for controlling for the organ damages such as spinal cord, brain, liver
It treats, and shows good therapeutic effect, and going back to the nest with the deviation of migratory direction is considered as weight present in stem-cell therapy
Want problem, it is therefore desirable to long-term lasting and noninvasive external tracking and monitoring means.It is extremely finely tissue that magnetic resonance imaging, which has,
Resolution ratio, spatial resolution, without free radiation and the features such as Non-Invasive, while can obtain transplanting surrounding tissue dissection and
Physiologic information, including the oedema and inflammation around transplanting stove.However with existing magnetic resonance resolution ratio, it is difficult directly to distinguish transplanting
Cell and host cell, this just needs to introduce certain image forming medium in advance in the cell, aobvious to cell to enhance magnetic resonance imaging
The sensibility shown.Previous research mostly uses SPIO nano particle to mark cell, and there are one for this labeling method
Intrinsic defect, i.e., as cell Proliferation passes on, content of the iron particle in cell can be gradually decreased, thus cannot achieve to shifting
Plant the long-term observation of cell.The imaging of magnetic resonance reporter gene overcomes this defect, and principle is that reporter gene is transferred to cell, leads to
Reporter gene continuous expression in the cell and Polyferric Sulfate (or other media) effect are crossed, cell is made to generate apparent magnetic resonance signal
Variation.Reporter gene imaging is a kind of noninvasive developing method carried out qualitatively and quantitatively in nucleus molecular level to biology, is led to
It is often then the carrier transfecting stem cells containing reporter gene implant, is situated between by the reporter protein of transfection cell expression
Lead the imaging that radioactive probe carries out Cellular tracking.Furthermore reporter gene provides turn that be non-invasive, visually detecting gene
A possibility that record and translation.Reporter gene is integrated into label cellular genome as carrier by slow virus, this insertion
The method advantage of gene is that reporter gene can copy in progeny cell with the proliferation passage of label cell, in gene expression
The consistency that parental cell and filial generation are maintained in level is capable of forming the consistent clonal population of gene expression dose, thus more
With the advantage in MRI signal comparison stability.
Iron ion is the important nutrient of cell growth, but excessive iron can cause cytotoxicity, keep the storage of iron ion
It deposits most important to cell with metabolic balance.Cell may be by excessive in ferritin or ferritin sample molecular memory cytoplasm
Iron come achieve the purpose that balance iron ion utilize and reduce cytotoxicity.At present have researcher by calculation biology prediction with
The full-length genome search and protein interaction experiment of drosophila propose the magnetic induction albumen being prevalent in animal
(Magnetoreceptor, the MagR) albumen belongs to iron-sulfur cluster assembling albumen, and there is iron to combine activity and apparent intrinsic magnetic moment,
It can be enriched with by magnetic field in laboratory and purifying obtains.It is highly important one kind in iron-sulfur cluster system that iron-sulfur cluster, which assembles albumen,
Iron-sulfur cluster assembles albumen, and there is internal iron to combine activity, while oxygen can inspire iron-sulfur cluster assembling albumen iron binding capacity, this
Kind can protect iron to generate not available iron hydroxide from the oxidation of oxygen under aerobic conditions in conjunction with activity.Iron-sulfur cluster
The biosynthesis of mitochondria iron-sulfur cluster under aerobic conditions may be participated in as a kind of molecular chaperones of iron by assembling albumen, and be prevented certainly
Lead to the generation of mitochondrial oxidation stress damage by iron doping.
Ferric citrate also known as iron ammonium citrate, are a kind of iron content, the complex salt of ammonia and citric acid, and ferric iron contains 5
Spin unpaired electron, and relaxation time of Hydrogen Proton in water is obviously shortened around it with very strong paramagnetic performance, concentration
Height can influence the effect of relaxation enhancing, and when low concentration is mainly shown as quickening longitudinal relaxation, and when high concentration is mainly shown as
Accelerate transverse relaxation.In vitro experiment, ferric citrate is a common exogenous iron additive, contains limited concentration model
Interior raising cell is enclosed to the absorption efficiency of iron ion, in order to form better signal contrast.Cell culture medium is added a certain amount of
Exogenous citrate iron ammonium, because ferric citrate must be reduced into inorganic ferrous salt as the organic molysite of trivalent in vivo could quilt
It absorbs.Ferric iron, when ferric iron enters in cell body, is turned by after cell active absorption by the reduction of cell membrane first
Become ferrous iron, and then is raised by magnetic induction albumen.
Summary of the invention
In order to solve the problems in the prior art, the present invention provides a kind of magnetic induction albumen as magnetic resonance imaging tracer
Application of the agent in stem cell tracer.Observe magnetic induction albumen reporter gene transfection cell overexpression situation and magnetic resonance at
As effect, the growing state and its migration situation of observation transfection cell.Magnetic induction albumen is one kind generally existing in organism
Ferritin is stored up, there is weak magnetic, endogenous NMR signal is can produce and compares and then MRI T1 longitudinal relaxation can be caused to believe
Number variation, shorten T1WI longitudinal relaxation time.The expression of magnetic induction albumen changes the changes in distribution of iron in the cell, to lure
The absorption of iron and the increase of cell iron content are led, prevents destruction of the active oxygen to cell at the same time.Magnetic induction albumen is really used
The intake to iron ion in surrounding tissue is relied primarily in the performance of internal Blue green algae, using ferric citrate as iron
Source promotes magnetic induction albumen to improve intracellular iron content to the intake of iron, and then can cause MRI T2 transverse-relaxation signals
Variation, it is significant to shorten T2WI lateral relaxation time, apparent low signal is shown on T2WI.
To achieve the goals above, the invention adopts the following technical scheme:
A kind of application of magnetic induction albumen as magnetic resonance imaging tracer in stem cell tracer.
Further, which includes: magnetic induction albumen inducing expression in stem cell, and magnetic induction albumen has internal iron
In conjunction with activity, free iron is raised, maintains the intracorporal iron stable state of machine, protection and prevention free iron are oxidized, in the work of externally-applied magnetic field
Under, Positive transfections stem cell is drawn towards magnetic direction, and stem cell directional migration makes stem cell obtain targeting transfer ability
Meanwhile magnetic resonance imaging carries out tracking stem cell from the overall process for being injected into differentiation.
Further, the magnetic induction albumen is by carrier transfecting stem cells, magnetic induction albumen and fluorescin to dry thin
Born of the same parents are marked jointly.
Further, the expression of the fluorescin detection magnetic induction albumen target gene, detects carrier package stem cell
The loss of target gene whether has occurred, and detects whether carrier is transferred to target gene the power expressed in target cell；Pass through
The fluorescent positive rate and fluorescence intensity of fluorescence microscope target gene cell, the expression of translation purpose gene.
Further, the magnetic induction albumen is Columba livia MagR (GeneBank MF134648.1).
Further, the stem cell includes embryonic stem cell and adult stem cell, and the adult stem cell fills between including
Matter stem cell, candidate stem cell.
Further, the free iron is ferrous ion.
Further, the carrier is virus, and the virus is slow virus (Lentivirus).
Further, the slow virus promoter is elongation factors EF1 α and cytomegalovirus CMV, and fluorescin is
eGFP。
Compared with prior art, of the invention that there is following beneficial effect:
(1) magnetic resonance imaging has the function of transplanting stem cell in noninvasive, repeatable tracer body, has good space point
Resolution, and imaging is not influenced by tissue depth.The indirect magnetic marker of stem cell makes cell express specific albumen by gene transfection
Matter enhances relaxation rate and contrast in tissue, improves T1 or T2 weighted imaging.Stem cell is expressed in the cell by transgenosis can be with
It is overexpressed by magnetic resonance signal or in the cell the albumen reacted with external source magnetic resonance imaging probe, to increase magnetic resonance
Imaging signal, to realize the vivo tracking to stem cell.With reporter gene be basic magnetic resonance imaging signal will not or seldom
It is influenced by stem cell division and passes to progeny cell in fission process, and the expression product of reporter gene can reflect
The function and vigor of stem cell.
(2) inactivation type viral vectors has many advantages, such as that biological safety is good, and transfection efficiency is high, is carried using it as gene delivery
Body, the cellular morphology indifference cultivated under the cell and normal morphology after transfection are almost not damaged to transfection target cell.Slow disease
Poison can infect division stage and nondividing phase cell, and foreign gene is effectively integrated on host chromosome, thus
Achieve the effect that persistence expresses aim sequence.
Detailed description of the invention
Fig. 1 is magnetic induction protein gene slow virus carrier expression system schematic diagram in the present invention；
Fig. 2 is that plasmid LV-EF1 α-MagR-CMV-eGFP-T2A-Puromycin digestion identifies that agarose is solidifying in the present invention
Gel electrophoresis figure；
Fig. 3 is that fluorescence microscopy microscopic observation recombinant slow virus transfection C57BL/6 mouse bone marrow cells mesenchyma is dry thin in the present invention
The luciferase expression situation of born of the same parents；
Fig. 4 is the melting curve of magnetic induction albumen and reference gene glyceraldehyde-3-phosphate dehydrogenase in the present invention.
Case is embodied
Below with reference to embodiment, the present invention will be further explained.
A kind of application of magnetic induction albumen as magnetic resonance imaging tracer in stem cell tracer, the application method are as follows:
Reporter gene magnetic induction albumen by slow virus carrier transfecting stem cells, using the distribution of magnetic resonance imaging monitoring transplanted cells,
The process of dynamic migration and integration with host, simultaneous selection fluorescin detect the table of magnetic induction albumen target gene
It reaches, whether detection Viral packaging cell has occurred the loss of target gene, and detects virus whether target gene to be transferred to target thin
The power expressed in born of the same parents.By the fluorescent positive rate and fluorescence intensity of fluorescence microscope target gene cell, i.e., indirectly
The expression of ground translation purpose gene.
Specifically, the application includes that reporter gene magnetic induction albumen inducing expression in stem cell, magnetic induction albumen has
Internal iron combination activity, is raised free iron (ferrous ion), maintains the intracorporal iron stable state of machine, protection and prevention ferrous ion
It is oxidized.
The magnetic induction albumen has following property: being distributed widely between different plant species, NCBI retrieval proves its level-one knot
The amino acid sequence of structure has well-conserved, and the function of homologous protein can be complementary and has internal iron knot between different plant species
Activity is closed, while oxygen can inspire magnetic induction albumen iron binding capacity, this combination activity can protect iron under aerobic conditions
Oxidation from oxygen generates not available iron hydroxide.Using transgene method, magnetic induction protein gene is led
Enter in stem cell, make its be overexpressed thus the ingestible more free iron of stem cell, magnetic resonance signal specific variations, to reach
Vivo tracking of the stem cell for a long time without invasion to after to transplanting.
Preferably, magnetic induction albumen is Columba livia MagR (GeneBank MF134648.1), magnetic induction albumen
Gene does not influence the activity of stem cell by carrier transfecting stem cells, magnetic induction protein gene.
Further, the magnetic induction albumen and fluorescin mark stem cell jointly, the fluorescin inspection
The expression of magnetic induction albumen target gene is surveyed, whether detection carrier package stem cell has occurred the loss of target gene, and detects
Whether carrier is transferred to target gene the power expressed in target cell；Pass through the fluorescence of fluorescence microscope target gene cell
Positive rate and fluorescence intensity, between translation purpose gene indirectly expression.
Preferably, fluorescin is enhanced green fluorescence protein (Enhanced green fluorescent
Protein, eGFP), it is made of 238 amino acid, by green fluorescent protein the 64th, 65 phenylalanine and serine point
It is not replaced as leucine and threonine, the green fluorescence under the excitation at 488nm wavelength of the eGFP after mutation is stronger, so as to it
It observes and detects and is more convenient sensitive.
Further, the stem cell is the cells of origin to form the various histoorgans of body, has self-renewing and more
To the function of differentiation, according to the stage of development and whether there is totipotency, stem cell is divided into embryonic stem cell and Cheng Tigan
Cell two major classes, in which: adult stem cell is a small amount of stem cell present in Various Tissues and organ, as mescenchymal stem cell,
Candidate stem cell etc., it is further preferable that the stem cell is C57BL/6 Marrow Mesenchymal Stem Cells.
Preferably, the carrier is virus, and the viral vectors selects slow virus (Lentivirus).It is further preferable that
The slow virus promoter is elongation factors EF1 α and cytomegalovirus CMV.
Embodiment 1, building slow virus carrier LV-EF1 α-MagR-CMV-eGFP-T2A-Puromycin:
Magnetic induction albumen reporter gene and enhanced green fluorescent protein gene are carried out total label to stem cell by the present invention,
Using double-promoter EF1 α and CMV, third generation recombinant slow virus packaging system, packaging plasmid, envelope plasmid, recombinant lentiviral disease are constructed
Cotransfection 293T cell after poisonous carrier plasmid is mixed with Opti-MEN finally obtains the slow virus for carrying target gene, qPCR method
Measure virus titer.The slow virus carrier building and identification for carrying MagR, eGFP and puromycin Puromycin, from
The cDNA sequence (Columba livia MagR, MF134648.1) of MagR gene is inquired in Genebank.
Prepare material: 3 helper plasmids (VSV-G, gag/pol, Rev), slow virus expression plasmid (pLV), 293T cell,
293T complete culture solution (DMEM+10%FBS), transfection reagent, HBSS；
1, packaging step:
1) day before transfection is inoculated with 293T cell into culture dish, and the cell to transfect the same day reaches 90% degrees of fusion and is
It is good；
2) the fresh 293T complete culture solution of 25mL is replaced before transfection；
3) application method for pressing transfection reagent, by 3 kinds of helper plasmids and pLV plasmid co-transfection 293T cell；
4) after 4-6 hours, 25mL 293T complete culture solution is replaced, sets and continues to cultivate in incubator；
5) after 48-72 hours, culture supernatant is collected into 50mL centrifuge tube, and 500g is centrifuged 5 minutes removal cell precipitations, returns
Receive supernatant；
6) supernatant is filtered with 0.45um filter, collects filtrate；
7) filtrate abandoned supernatant with 50000g high speed centrifugation 2 hours.It is as dense with the abundant lytic virus precipitating of appropriate HBSS
Contracting slow virus.
2, digestion is identified
By the LV-EF1 α-MagR-CMV-eGFP-T2A-Puromycin plasmid of purifying, limited using Sacl restriction enzyme, Xmnl
Enzyme and Ncol restriction enzyme processed carries out specific site digestion and carries out 1% gel electrophoresis identification；
3, titre detects
1) QPCR standard items are made: gene (ALB) segment of intracellular known copy number being cloned into carrier and extracts matter
Grain, as standard items 1；Standard items 2 are slow virus expression plasmid.
2) transduction the previous day is inoculated with 293T cell into 6 orifice plates or 12 orifice plates；
3) before transduceing, the cell for digesting a hole carries out cell count, obtains its cell concentration (A)；
4) viral dilution certain multiple (B) transduces 293T cell (volume V mL), and polybrene is added and promotes infection；
5) it after 48 hours, collects cell and extracts its genomic DNA；
6) QPCR: using two standard curves, obtains the ALB copy number and WPRE copy number of DNA sample respectively, calculates
The copy number C of virus sequence in the cell of average each sample；
7) virus titer (TU/mL)=(target cell (A) × extension rate B × C)/virus volume (V)
As shown in Figure 1, the slow related viral vectors expression system schematic diagram of magnetic induction protein gene, comprising EF1 α, (people extends
The factor) promoter, cytomegalovirus CMV, self cleavage T2A, magnetic induction albumen MagR, enhanced green fluorescence protein eGFP and fast
Purine mycin Puromycin.
As shown in Fig. 2, recombinant virus carries out Sacl restriction enzyme/Xmnl restriction enzyme/Ncol restriction enzyme digestion identification, digestion is produced
Object carries out 1.0% gel electrophoresis, and Scal limits digestion and obtains 3700bp, 2300bp, 2000bp as the result is shown, 1600bp and
200bp, Xmnl limitation digestion obtain 4400bp, 2500bp, 1600bp and 1100bp, and Ncol limitation digestion obtains 6100bp,
2000bp, 800bp, 600bp and 200bp, the above results show successfully to construct the LV-EF1 α-MagR-CMV- containing MagR
EGFP-T2A-Puromycin slow virus carrier.
As shown in table 1, virus titer measurement result, the slow disease collected after being concentrated and purified using the detection of absolute quantitation PCR method
Poison, LV-EF1 α-MagR-CMV-eGFP-T2A-Puromycin virus titer are 2.40 × 108TU/mL, LV-CMV-eGFP-
T2A-Puromycin virus titer is 6.92 × 108TU/mL。
Table 1: virus titer measurement result
Embodiment 2, building magnetic induction PROTEIN C 57BL/6 Marrow Mesenchymal Stem Cells surely turn strain:
1, virus transfection C57BL/6 Marrow Mesenchymal Stem Cells,
1) cell prepares: by cell inoculation to 6 orifice plates, should guarantee as single cell suspension as far as possible and digestible degree is suitble to, connect
Kind cell concentration is about 2.5 × 105-3.5×105The hole cells/, every hole 2mL complete medium, cell should be at good before passing on
Cell is placed in 37 DEG C, 5%CO by growth conditions2, 95% relative humidity incubator in cultivate 18-24h；
2) viral transduction: the transfection same day takes out cell and is observed, and needs to ensure that cell density is suitable for that (convergence degree is before transduction
30-50%), in good condition, a wherein hole cell is digested with pancreatin and is counted, and general every hole cell number is 3 × 105；
3) culture medium is drawn, 1ml fresh complete medium is changed to, is put into incubator before transduction and is balanced；
4) melt slow virus, being put to ice bath or 4 DEG C melts it, needs to keep its guarantee low in merging ice chest after taking-up
Temperature；
5) it after virus is melted, slightly inhales and plays mixing, according to virus titer and the most suitable infection multiplicity of cell, after sufficiently shaking up, often
Hole can be added polybrene (ultimate density should be 5 μ g/mL) and improve transfection efficiency；
6) incubator is put into be cultivated, after 6-15 hours observe cell state (such as visible cell has overt toxicity reaction,
Transduction time can be shortened was advisable at 6-8 hours), and it is changed to fresh complete medium, it takes pictures after the 48h that transduces, Yu Ying
Viewed under light microscopy slow-virus transfection effect；
2, real-time fluorescence quantitative PCR detection C57BL/6 Marrow Mesenchymal Stem Cells magnetic induction albumen-mRNA expression:
1) Total RNAs extraction carries out the extraction of total serum IgE using TIANGEN (Tiangeng) total RNA from animal tissues extracts kit,
Period carries out DNA and handles；
2) total serum IgE reverse transcription utilizes TaKaRa (precious bioengineering (Dalian) Co., Ltd) PrimeScriptTMRT
The reverse transcription of reagent Kit with gDNA Eraser (Perfect Real Time) kit progress total serum IgE；
3) quantitative fluorescent PCR is contaminated using glyceraldehyde-3-phosphate dehydrogenase GAPDH gene as internal reference using SYBR Green I
Material method carries out relative quantitative assay to the expression of target gene in the sample, and each sample does three parallel, experiment primers
With qPCR shown in reaction system such as table 2.
Table 2
As shown in figure 3, after recombinant slow virus LV-EF1 α-MagR-CMV-eGFP-T2A-Puromycin is transfected 48 hours,
In the lower 100 times of observations of inverted fluorescence microscope, there is apparent green fluorescence into the cell, with the unloaded slow virus LV-CMV- of transfection
EGFP-T2A-Puromycin is compared, and transfection efficiency is suitable, and about 80% cell by slow-virus infection and generates fluorescence, is shown
Slow virus Successful transfection enters Marrow Mesenchymal Stem Cells, surely turns strain and constructs and screen success.The figure of the first row in Fig. 3 are as follows:
The figure of second row in zero load group transfection LV-CMV-eGFP-T2A-Puromycin, Fig. 3 are as follows: experimental group transfects LV-EF1 α-MagR-
CMV-eGFP-T2A-Puromycin。
As shown in figure 4, passing through the visible magnetic induction albumen of solubility curve figure that instrument software automatically generates and reference gene
Melting curve is simple spike, has no other wave crests, shows that this is established the amplified production of qPCR method and has specificity and determine
Amount is accurate.
As shown in table 3, cycle-index C in the present inventionTRNA real time fluorescent quantitative is extracted in the measurement of value and melting temperature Tm
QPCR detects the expression of MagR-mRNA, with the correction of hGAPDH mRNA level in-site, tests eGFP empty plasmid as the result is shown and does not almost have
There is MagR-mRNA expression, illustrate that the virus particle of experimental group MagR constructs successfully, and successfully pack in cell, has thin
The ability of successful expression MagR intracellular.
Table 3: cycle-index C in the present inventionTThe measurement of value and melting temperature Tm
| Sample ID | MagR TmMelting temperature | hGAPDH TmMelting temperature |
| Experimental group | 79.5126℃ | 85.9344℃ |
| Unloaded group | - | 85.1093℃ |
| Sample ID | MagR CT cycle-index | hGAPDH CT cycle-index |
| Experimental group | 15.758 | 13.163 |
| Unloaded group | - | 13.163 |
The sequence table of the magnetic induction albumen is as follows:
Columba livia MagR (GeneBank MF134648.1) cDNA:
1ATGGCC TCGTCG GCCTCG TCCGTG GTGAGG GCCACG GTGCGC GCCGTC AGCAAG
CGGAAG
61ATCCAG GCTACG CGCGCC GCCCTC ACCCTG ACCCCA TCAGCT GTTCAG AAGATA
AAAGAG
121CTTCTT AAAGAT AAACCT GAGCAT GTAGGC GTGAAA GTAGGT GTTCGC ACAAGA
GGATGC
181AATGGA CTTTCT TACACA TTAGAA TATACA AAATCA AAAGGA GACTCT GATGAA
GAAGTA
241GTTCAA GATGGG GTTAGA GTGTTT ATTGAG AAGAAG GCACAG CTGACG CTTTTA
GGCACT
301GAAATG GACTAT GTAGAA GACAAA CTGTCC AGTGAA TTTGTC TTCAAT AATCCA
AACATC
361AAAGGA ACATGT GGCTGT GGAGAA AGCTTT AACATC TGA
The above is only a preferred embodiment of the present invention, it should be pointed out that: for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Sequence table
<110>Southeast China University
<120>application of the magnetic induction albumen as magnetic resonance imaging tracer in stem cell tracer
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 399
<212> DNA
<213>magnetic induction albumen (MagR)
<400> 1
atggcctcgt cggcctcgtc cgtggtgagg gccacggtgc gcgccgtcag caagcggaag 60
atccaggcta cgcgcgccgc cctcaccctg accccatcag ctgttcagaa gataaaagag 120
cttcttaaag ataaacctga gcatgtaggc gtgaaagtag gtgttcgcac aagaggatgc 180
aatggacttt cttacacatt agaatataca aaatcaaaag gagactctga tgaagaagta 240
gttcaagatg gggttagagt gtttattgag aagaaggcac agctgacgct tttaggcact 300
gaaatggact atgtagaaga caaactgtcc agtgaatttg tcttcaataa tccaaacatc 360
aaaggaacat gtggctgtgg agaaagcttt aacatctga 399
Claims (9)
1. a kind of application of magnetic induction albumen as magnetic resonance imaging tracer in stem cell tracer.
2. application according to claim 1, which is characterized in that magnetic induction albumen inducing expression in stem cell, magnetic induction
Albumen has internal iron combination activity, raises free iron, maintains the intracorporal iron stable state of machine, and protection and prevention free iron are oxidized.
3. application according to claim 1, which is characterized in that the magnetic induction albumen is by carrier transfecting stem cells, magnetic
Induction albumen and fluorescin mark stem cell jointly.
4. application according to claim 3, which is characterized in that the fluorescin detection magnetic induction albumen target gene
Expression, whether detection carrier package stem cell has occurred the loss of target gene, and detects whether carrier is transferred to target gene
The power expressed in target cell；By the fluorescent positive rate and fluorescence intensity of fluorescence microscope target gene cell, turn over
Translate the expression of target gene.
5. application according to claim 2, which is characterized in that the magnetic induction albumen is Columba livia MagR
(GeneBank MF134648.1)。
6. application according to claim 2, which is characterized in that the stem cell includes embryonic stem cell and thin at soma
Born of the same parents, the adult stem cell include mescenchymal stem cell, candidate stem cell.
7. application according to claim 2, which is characterized in that the free iron is ferrous ion.
8. application according to claim 3, which is characterized in that the carrier is virus, and the virus is slow virus
(Lentivirus)。
9. application according to claim 8, which is characterized in that the slow virus promoter be elongation factors EF1 α with it is big and small
Cellular virus CMV, fluorescin eGFP.
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| CN114908055A (en) * | 2022-05-13 | 2022-08-16 | 东南大学 | Stem cell for over-expressing Magr to regulate iron homeostasis in vivo and preparation method thereof |
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| CN114908055B (en) * | 2022-05-13 | 2024-03-12 | 东南大学 | Stem cells for over-expressing Magr to regulate in-vivo iron homeostasis and preparation method thereof |
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