Chromosomal FISH-TSA - Revision history

imported>Ksnyder at 22:43, 10 January 2011

2011-01-10T22:43:45Z

← Older revision		Revision as of 22:43, 10 January 2011
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	'''Protocol:'''		'''Protocol:'''

	~~'''A. Preparation of labeled cDNA probe'''~~		* [[Image:Krylov_FISH-TSA_protocol.pdf \|Data mapping document]]

	~~We always use the cDNA probe of approx. 1000 bp in length depending on type of cDNA. Any repete sequences in the probe could make a strong background using TSA.~~

	~~- prepare at least 2 μg of amplificate of cDNA from reverse transcription reaction by PCR and specific primers~~

	~~- purify the amplificate through agarose gel and column of Gel extraction kit (QIAGEN)~~

	~~- label this cDNA by Digoxigenine~~-11 dUTP alkali stable (Roche) and Deca Label Labelling kit (Fermentas) – labelling by random primers. We always label 1μg of cDNA for 20 hours. After this time we purify the probe with column of gel extraction kit (QIAGEN) without using an agarose. Just mix probe with binding buffer. Elution is done with 50 μl of sterile water. We always use 2 μl of probe in 50 μl reaction per one slide.

imported>Xenbase: moved FISH-TSA to Chromosomal FISH-TSA

2011-01-10T17:19:42Z

moved FISH-TSA to Chromosomal FISH-TSA

← Older revision	Revision as of 17:19, 10 January 2011
(No difference)

136.159.234.129 at 18:50, 3 January 2011

2011-01-03T18:50:58Z

← Older revision		Revision as of 18:50, 3 January 2011
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	Following protocol is designed for the visualization of short cDNA sequences with the minimal lenght of 900-1000 bp.		Following protocol is designed for the visualization of short cDNA sequences with the minimal lenght of 900-1000 bp.

	'''~~P R O T O C O L~~:'''		'''Protocol:'''

	'''A. Preparation of labeled cDNA probe'''		'''A. Preparation of labeled cDNA probe'''

	We always use the cDNA probe of approx. 1000 bp in length depending on type of cDNA. Any ~~repetition~~ could make a strong background using TSA.		We always use the cDNA probe of approx. 1000 bp in length depending on type of cDNA. Any repete sequences in the probe could make a strong background using TSA.

	- prepare at least 2 μg of amplificate of cDNA from reverse transcription reaction by PCR and specific primers		- prepare at least 2 μg of amplificate of cDNA from reverse transcription reaction by PCR and specific primers

195.113.59.254 at 10:41, 8 December 2010

2010-12-08T10:41:34Z

← Older revision		Revision as of 10:41, 8 December 2010
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	Krylov V., Tlapakova T., Macha J. (2007) Localization of Mdh2 single copy gene on Xenopus tropicalis chromosomes by FISH-TSA technique. Cytogenet. Genome Res. 116:110-112. [http://www.ncbi.nlm.nih.gov/pubmed/17268187].		Krylov V., Tlapakova T., Macha J. (2007) Localization of Mdh2 single copy gene on Xenopus tropicalis chromosomes by FISH-TSA technique. Cytogenet. Genome Res. 116:110-112. [http://www.ncbi.nlm.nih.gov/pubmed/17268187].

			Following protocol is designed for the visualization of short cDNA sequences with the minimal lenght of 900-1000 bp.

			'''P R O T O C O L:'''

			'''A. Preparation of labeled cDNA probe'''

			We always use the cDNA probe of approx. 1000 bp in length depending on type of cDNA. Any repetition could make a strong background using TSA.

			- prepare at least 2 μg of amplificate of cDNA from reverse transcription reaction by PCR and specific primers

			- purify the amplificate through agarose gel and column of Gel extraction kit (QIAGEN)

			- label this cDNA by Digoxigenine-11 dUTP alkali stable (Roche) and Deca Label Labelling kit (Fermentas) – labelling by random primers. We always label 1μg of cDNA for 20 hours. After this time we purify the probe with column of gel extraction kit (QIAGEN) without using an agarose. Just mix probe with binding buffer. Elution is done with 50 μl of sterile water. We always use 2 μl of probe in 50 μl reaction per one slide.

195.113.59.254 at 10:25, 8 December 2010

2010-12-08T10:25:12Z

← Older revision		Revision as of 10:25, 8 December 2010
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	Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res. 2003;103(1-2):169-172. [http://www.ncbi.nlm.nih.gov/pubmed/15004482].		Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res. 2003;103(1-2):169-172. [http://www.ncbi.nlm.nih.gov/pubmed/15004482].

			Tlapakova T., Krylov V., Macha J. (2005) Localization, structure and polymorphism of two paralogous Xenopus laevis mitochondrial malate dehydrogenase genes. Chromosome res. 13(7):699-706. [http://www.ncbi.nlm.nih.gov/pubmed/16235119].

			Krylov V., Tlapakova T., Macha J. (2007) Localization of Mdh2 single copy gene on Xenopus tropicalis chromosomes by FISH-TSA technique. Cytogenet. Genome Res. 116:110-112. [http://www.ncbi.nlm.nih.gov/pubmed/17268187].

195.113.59.254 at 10:21, 8 December 2010

2010-12-08T10:21:17Z

← Older revision		Revision as of 10:21, 8 December 2010
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	'''Literature:'''		'''Literature:'''

	Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res. 2003;103(1-2):169-172. [http://www.ncbi.nlm.nih.gov/pubmed/15004482].		Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res. 2003;103(1-2):169-172. [http://www.ncbi.nlm.nih.gov/pubmed/15004482].

195.113.59.254 at 10:20, 8 December 2010

2010-12-08T10:20:46Z

← Older revision		Revision as of 10:20, 8 December 2010
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	Literature:		'''Literature:'''

	Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res. 2003;103(1-2):169-172. [http://www.ncbi.nlm.nih.gov/pubmed/15004482].		Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res. 2003;103(1-2):169-172. [http://www.ncbi.nlm.nih.gov/pubmed/15004482].

195.113.59.254: Created page with "Literature: Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res..."

2010-12-08T10:19:09Z

Created page with "Literature: Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res..."

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Literature:

Krylov V., Macha J., Tlapakova T., Takac M., Jonak J. (2003) The c-SRC1 gene visualized by in situ hybridization on Xenopus laevis chromosomes. Cytogenet. Genome Res. 2003;103(1-2):169-172. [http://www.ncbi.nlm.nih.gov/pubmed/15004482].