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1. Harvest cells from a culture that is at or near peak density by centrifugation at 800 x g for 5 min. The cells grown in a medium containing agar are concentrated by centrifugation, a solid pellet does not form. The soft pellet is re-suspended to desired cell concentration with agar-free supernatant.
2. Adjust the concentration of cells to 2 x 10⁶ - 10⁷/mL in fresh medium.
3. While cells are centrifuging prepare a 10% (v/v) solution of sterile DMSO in fresh medium.
   1. Add 1.0 mL of DMSO to an ice cold 20 x 150 mm screw-capped test tube;
   2. Place the tube on ice and allow the DMSO to solidify (~5 min) and then add 9.0 mL of ice cold medium;
   3. Invert several times to dissolve the DMSO;
   4. Allow to warm to room temperature.
4. Mix the cell preparation and the DMSO in equal portions. Thus, the final concentration will be 10⁶ - 10⁷ cells/mL and 5% (v/v) DMSO. The time from the mixing of the cell preparation and DMSO stock solution before the freezing process is begun should no less than 15 min and no longer than 30 min.
5. Dispense in 0.5 mL aliquots into 1.0 - 2.0 mL sterile plastic screw-capped cryules (special plastic vials for cryopreservation). 
6. Place the vials in a controlled rate freezing unit.  Use the following cooling cycle: From room temperature cool at -10°C/min to the heat of fusion; from the heat of fusion to -40°C, cool at -1°C/min.  At -40°C plunge into liquid nitrogen.  The cooling cycle should be initiated no less than 15 and no more than 30 minutes after the addition of DMSO to the cell preparation.
7. The frozen preparations should be stored in either the vapor or liquid phase of a nitrogen refrigerator. Frozen preparations stored below -130°C are stabile indefinitely. Those stored at temperatures above -130°C are progressively less stabile as the storage temperature is elevated. Vials should not be stored above -55°C.
8. To establish a culture from the frozen state place an ampule in a water bath set at 35°C. Immerse the vial just to a level just above the surface of the frozen material. Do not agitate the vial.
9. Immediately after thawing, do not leave in the water bath, aseptically remove the contents of the ampule and inoculate a 16 x 125 mm screw-capped test tube containing 13 mL of ATCC medium 2692
10. Incubate the culture on a 15° horizontal slant at 35°C.

Alderete JF, Garza GE. Identification and properties of Trichomonas vaginalis proteins involved in cytadherence. Infect. Immun. 56: 28-33, 1988. PubMed: 3257206

Diamond LS. Axenic cultivation of Trichomonas tenax, the oral flagellate of man. I. Establishment of cultures. J. Protozool. 9: 442-444, 1962.

Gannon JT, Linke HA. Growth studies on xenic cultures of Entamoeba gingivalis using established media. Int. J. Parasitol. 19: 835-838, 1989. PubMed: 2635159

Poirier TP, et al. . Trans. Am. Microsc. Soc. 109: 342-351, 1990.

Yamamoto A, et al. Nucleotide sequence of the SrRNA gene of Entamoeba gingivalis: applications for construction of a species-specific DNA probe and phylogenetic analysis. Microbiol. Immunol. 39: 185-192, 1995. PubMed: 7603363

Fukura K, et al. Nucleotide Sequence of the SrRNA Gene and Phylogenetic Analysis of Trichomonas tenax. Microbiol. Immunol. 40: 183-188, 1996. PubMed: 8934671

Felleisen RS, et al. Detection of Tritrichomonas foetus by PCR and DNA enzyme immunoassay based on rRNA gene unit sequences. J. Clin. Microbiol. 36: 513-519, 1998. PubMed: 9466768

Felleisen RS. Comparative sequence analysis of 5.8S rRNA genes and internal transcribed spacer (ITS) regions of trichomonadid protozoa. Parasitology 115: 111-119, 1997. PubMed: 10190167

Madico G, et al. Diagnosis of Trichomonas vaginalis infection by PCR using vaginal swab samples. J. Clin. Microbiol. 36: 3205-3210, 1998. PubMed: 9774566

Yamamoto A, et al. Phylogenetic position of the mitochondrion-lacking protozoan Trichomonas tenax, based on amino acid sequences of elongation factors 1alpha and 2. J. Mol. Evol. 44: 98-105, 1997. PubMed: 9010141

Nagao E, et al. Two distinct hemolysins in Trichomonas tenax ATCC 30207. Oral Microbiol. Immunol. 15: 355-359, 2000. PubMed: 11154431

Gerbod D, et al. Phylogenetic relationships of class II fumarase genes from trichomonad species. Mol. Biol. Evol. 18: 1574-1584, 2001. PubMed: 11470849

Nucleotide (GenBank) : D49495 Trichomonas tenax gene for SrRNA.

Nucleotide (GenBank) : D78480 elongation factor 2, partial coding sequence

Nucleotide (GenBank) : D78481 elongation factor 2, partial coding sequence

Nucleotide (GenBank) : D78479 Trichomonas tenax gene for elongation factor 1 alpha, partial cds.

Nucleotide (GenBank) : U86615 putative rRNA small subunit gene, partial sequence, putative internal transcribed spacer 1, putative 5.8S rRNA gene, and putative internal transcribed spacer 2, complete sequence, and putative rRNA large subunit gene, partial sequence