Spolpitiyaarachchige at 15:51, 11 June 2013

2013-06-11T15:51:08Z

Spolpitiyaarachchige at 15:50, 11 June 2013

2013-06-11T15:50:42Z

Spolpitiyaarachchige: Created page with "PCR primers for insertion of Genes between Junctions C and D 1. Conduct Inverse PCR using scaffold with promoters and C dogs and J-GGA_C_rev and J-GGA_D_for. 2. Conduct PCR ..."

2013-06-11T15:49:53Z

Created page with "PCR primers for insertion of Genes between Junctions C and D 1. Conduct Inverse PCR using scaffold with promoters and C dogs and J-GGA_C_rev and J-GGA_D_for. 2. Conduct PCR ..."

New page

PCR primers for insertion of Genes between Junctions C and D

1. Conduct Inverse PCR using scaffold with promoters and C dogs and J-GGA_C_rev and J-GGA_D_for.

2. Conduct PCR of eCDM8 gene and RFP gene templates with primers below.

3. Conduct J-GGA with products from 1 and 2 above.

Gene primer considerations.

1. Four extra bases on 5’ end (eg. GCAT)

2. BsaI site directed toward the gene

3. Sticky end of 5’ CATC 3’ for Junction C on For primer

4. Sticky end of 5’ TCCG 3’ for Junction D on Rev primer

5. 18 nt of gene coding sequence on each primer

Primer designs for the eCDM8 gene

Forward ECDM8 gcatggtctcacatcaaagaaatgccgcagccg

Rev ECDM8 ccagggtctcatccgttattaacctgcccaaac

5’ gttTgggcaggttaataa 3’

3’ caaacccgtccaattatt 5’

>BBa_J100100 Part-only sequence (3204 bp)
gaattcgcggccgcttctagagggtctcgaatgaccatcaaagaaatgccgcagccgaaaacctttggtgaactgaaaaatctgccgctgctgaataccg

ataaaccggttcatgcactgatgaaaattgcagatgaactgggcgagatctttaaattcgaagcaccgggtagcgttacccgttatctgagcagccagcg

tctgattaaagaagcatgcgacgaaagccgctttgataaaaatctgattcagtggctgaaattcatccgcgattttctgggtgatggtctgagcaccagc

tggacccatgaaaaaaactggaaaaaagcccataatattctgctgccgagctttagccagcaggcaatgaaaggttatcatgcaatgatggttgatattg

ccgttcagctggttcagaaatgggaacgtctgaatgcagatgaacatattgaagttccggaagatatgacccgtctgaccctggataccattggtctgtg

tggttttaactatcgcttcaacagcttttatcgtgatcagccgcatccgtttgttaccagcatggttcgtgcactggatgaagcagtgaataaatggcag

cgtgcaaatccggatgatccggcatatgatgaaaataaacgtcagttccaagaggatattaaagtgatgaatgacctggtggacaaaattatcgcagatc

gtaaagcaagcggtgaacagagtgatgatctgctgacccatatgctgaatggtaaagatccggaaaccggtgaaccgctggatgatgaaaacattcgtta

tcagattattaccttcctgattgccggtcatgaaaccaccagtggtctgctgagctttgcactgtattttctggttaaaaatccgcacgttctgcaaaaa

gcagccgaagaagcagcacgcgttctggttgatccggttccgagctataaacaggttaaacagctgaaatatgtgggcatggttctgaatgaagcactgc

gtctgtggccgacctttccgtggtttagcctgtatgcaaaagaagataccgttctgggaggtgaatatccgctggaaaaaggtgatgaactgatggttct

gattccgcagctgcatcgtgataaaaccatttggggtgatgatgtggaagaatttcgtccggaacgttttgaaaatccgagcgcaattccgcagcatgcc

tttaaaccgtttggtaatggtcagcgtgcatgtattggtcagcagtttgcactgcatgaagccaccctggttctgggtatgatgctgaaacattttgatt

ttgaggaccacaccaactatgagctggatattaaaggcaccctgaccctgaaaccggaaggttttgttgttaaagccaaaagcaaaaaaatcccgctggg

tggtattccgagcccgagcaccgaacagagcgcaaaaaaagttcgtaaaaaagccgaaaacgcccataatacaccgctgctggttctgtatggtagcaat

atgggtacagccgaaggcaccgcacgtgatctggcagatattgcaatgagcaaaggttttgcaccgcaggttgcaaccctggatagccatgcaggtaatc

tgcctcgtgaaggtgcagttctgattgttaccgcaagctataatggtcatccgcctgataatgcaaaacagtttgttgattggctggatcaggcaagcgc

agatgaagttaaaggtgttcgttatagcgtttttggttgcggtgataaaaactgggcaaccacctatcagaaagttccggcatttattgatgaaaccctg

gcagcaaaaggtgcagaaaatattgcagatcgtggtgaaaccgatgccagtgatgattttgaaggcacctatgaagaatggcgtgaacatatgtggtcag

atgttgcagcctattttaacctggatatcgaaaacagcgaggacaataaaagcaccctgagcctgcaatttgtggatagcgcagcagatatgccgctggc

aaaaatgcatggtgcatttagcaccaatgttgttgcaagcaaagaactgcaacagcctggtagcgcacgtagcacccgtcatctggaaattgaactgccg

aaagaagcaagctatcaagagggtgatcatctgggtgttattccgcgtaattatgaaggtattgttaatcgtgttaccgcacgttttggtctggatgcaa

gccagcagattcgtctggaagcagaagaagaaaaactggcacatctgccgctggccaaaaccgttagcgttgaagaactgctgcaatatgttgaactgca

agatccggttacccgtacccagctgcgtgcaatggcagcaaaaaccgtttgtccgcctcataaagttgaactggaagcactgctggagaaacaggcatat

aaagaacaggttctggcaaaacgtctgaccatgctggaactgctggaaaaatatccggcatgcgaaatgaaattcagcgaatttattgcactgctgccga

gtattcgtccgcgttattatagcattagcagcagtccgcgtgttgatgaaaaacaggccagcattaccgttagcgtggttagcggtgaagcatggtcagg

ttatggtgaatataaaggtattgccagcaattatctggccgaactgcaagaaggtgataccattacctgttttattagcacaccgcagagcgaatttacc

ctgccgaaagatcctgaaacaccgctgattatggttggtccgggtacaggtgttgcaccgtttcgtggttttgttcaggcacgtaaacaactgaaagaac

agggtcagagcctgggtgaagcacatctgtattttggttgtcgtagtccgcatgaggattatctgtatcaagaagaactggaaaacgcacagagcgaagg

tattattaccctgcataccgcatttagccgtatgccgaatcagccgaaaacatatgttcagcatgttatggaacaggatggcaaaaaactgattgaactg

ctggatcagggtgcccatttctatatttgtggtgatggtagccagatggcaccggcagttgaagcaaccctgatgaaaagctatgcagatgttcatcagg

ttagcgaagcagatgcacgtctgtggctgcaacagctggaagaaaaaggtcgttatgcaaaagatgtttgggcaggttaataatactagtagcggccgct
gcag

Primer designs for the RFP gene

RFP Forward gcatggtctcgcatcgcttcctccgaagacgtt

RFP Rev ccagggtctcatccgttattaagcaccggtgga

5’ Tccaccggtgcttaataa 3’

3’ Aggtggccacgaattatt 5’

RFP Gene Sequence

>BBa_E1010 Part-only sequence (706 bp)

atggcttcctccgaagacgttatcaaagagttcatgcgtttcaaagttcgtatggaaggttccgttaacggtcacgagttcgaaatcgaaggtgaaggtg

aaggtcgtccgtacgaaggtacccagaccgctaaactgaaagttaccaaaggtggtccgctgccgttcgcttgggacatcctgtccccgcagttccagta

cggttccaaagcttacgttaaacacccggctgacatcccggactacctgaaactgtccttcccggaaggtttcaaatgggaacgtgttatgaacttcgaa

gacggtggtgttgttaccgttacccaggactcctccctgcaagacggtgagttcatctacaaagttaaactgcgtggtaccaacttcccgtccgacggtc

cggttatgcagaaaaaaaccatgggttgggaagcttccaccgaacgtatgtacccggaagacggtgctctgaaaggtgaaatcaaaatgcgtctgaaact

gaaagacggtggtcactacgacgctgaagttaaaaccacctacatggctaaaaaaccggttcagctgccgggtgcttacaaaaccgacatcaaactggac

atcacctcccacaacgaagactacaccatcgttgaacagtacgaacgtgctgaaggtcgtcactccaccggtgcttaataa

← Older revision		Revision as of 15:51, 11 June 2013
Line 34:		Line 34:

	>BBa_J100100 Part-only sequence (3204 bp)		>BBa_J100100 Part-only sequence (3204 bp)
		+
	gaattcgcggccgcttctagagggtctcgaatgaccatcaaagaaatgccgcagccgaaaacctttggtgaactgaaaaatctgccgctgctgaataccg		gaattcgcggccgcttctagagggtctcgaatgaccatcaaagaaatgccgcagccgaaaacctttggtgaactgaaaaatctgccgctgctgaataccg

@@ Line 1: / Line 1: @@
 PCR primers for insertion of Genes between Junctions C and D
 .	Conduct Inverse PCR using scaffold with promoters and C dogs and J-GGA_C_rev and J-GGA_D_for.
@@ Line 6: / Line 7: @@
 .	Conduct J-GGA with products from 1 and 2 above.
 Gene primer considerations.
@@ Line 18: / Line 20: @@
 .	18 nt of gene coding sequence on each primer
 Primer designs for the eCDM8 gene
 Forward ECDM8 gcatggtctcacatcaaagaaatgccgcagccg
@@ Line 96: / Line 100: @@
 Primer designs for the RFP gene
 RFP  Forward      gcatggtctcgcatcgcttcctccgaagacgtt
 RFP Rev                ccagggtctcatccgttattaagcaccggtgga
 ’ Tccaccggtgcttaataa 3’
 ’ Aggtggccacgaattatt 5’
 RFP Gene Sequence

Primer Design for the eCDM8 and RFP - Revision history

Spolpitiyaarachchige at 15:51, 11 June 2013

Spolpitiyaarachchige at 15:50, 11 June 2013

Spolpitiyaarachchige: Created page with "PCR primers for insertion of Genes between Junctions C and D 1. Conduct Inverse PCR using scaffold with promoters and C dogs and J-GGA_C_rev and J-GGA_D_for. 2. Conduct PCR ..."