User:Stuart McKellar/Notebook/Bird Sex Testing/2013/01/04

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(Autocreate 2013/01/04 Entry for User:Stuart_McKellar/Notebook/Bird_Sex_Testing)
Current revision (18:16, 8 January 2013) (view source)
(Temps and new primers)
 
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==Entry title==
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==Reconciling information and thoughts after break==
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* Insert content here...
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I did some reading and thinking and research during the xmas break. This is the result.
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=Housekeeping Genes=
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Finally found some housekeeping primers:
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18S-F908 18S a AGCGAAAGCATTTGCCAAGA 401 This study (SVB)
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18S-R1309 18S a AGTCTCGTTCGTTATCGGAATT This study (SVB)
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(from http://wwwnc.cdc.gov/eid/article/11/5/05-0211-t1.htm)
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=Temps and new primers=
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P2/P8 primers work best at 60C, confirming my lab work.
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(from http://www.academia.edu/1460726/Sex_Identification_of_Some_Pet_Birds_by_Polymerase_Chain_Reaction-Based_Methods )
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also, this paper suggests using some new primers and was getting results very similar to my ones. Going to try and order NP an MP primers. See this paper for what a positive and negative result looks like using P2/P8 primers.
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NP primer:
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5’-GAG AAA CTG TGC AAAACA G-3’
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MP primer:
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5’-AGT CAC TAT CAG ATC CGG AA-3’
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Also this page suggests using 1μL of 10μM primer to give final concentration of 0.5μM in 20μL. Maybe I should start running the reactions at 20ul instead.
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Mushy primers:
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ITS1 5'-TCCGTAGGTGAACCTGCGG-3' forward all
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ITS4 5'-TCCTCCGCTTATTGATATGC-3' reverse all
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Reconciling information and thoughts after break

I did some reading and thinking and research during the xmas break. This is the result.

Housekeeping Genes

Finally found some housekeeping primers: 18S-F908 18S a AGCGAAAGCATTTGCCAAGA 401 This study (SVB) 18S-R1309 18S a AGTCTCGTTCGTTATCGGAATT This study (SVB)

(from http://wwwnc.cdc.gov/eid/article/11/5/05-0211-t1.htm)

Temps and new primers

P2/P8 primers work best at 60C, confirming my lab work. (from http://www.academia.edu/1460726/Sex_Identification_of_Some_Pet_Birds_by_Polymerase_Chain_Reaction-Based_Methods )

also, this paper suggests using some new primers and was getting results very similar to my ones. Going to try and order NP an MP primers. See this paper for what a positive and negative result looks like using P2/P8 primers.

NP primer: 5’-GAG AAA CTG TGC AAAACA G-3’

MP primer: 5’-AGT CAC TAT CAG ATC CGG AA-3’

Also this page suggests using 1μL of 10μM primer to give final concentration of 0.5μM in 20μL. Maybe I should start running the reactions at 20ul instead.


Mushy primers: ITS1 5'-TCCGTAGGTGAACCTGCGG-3' forward all ITS4 5'-TCCTCCGCTTATTGATATGC-3' reverse all

EDIT THESE PAGES TO SEE NICE FORMATTING


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