Nucleic Acids Research, Vol. 18, No. 18 5563
.. 1990 Oxford University Press
of feline tumor necrosis factor alpha
Gene sequence
Royal A.McGraw*, Bradford W.Coffee, Cynthia M.Ottol, Roman T.Drews and Clarence A.Rawlings1 Departments of Physiology and Pharmacology and 'Small Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA EMBL accession
Submitted August 3, 1990
This work was supported by the University of Georgia Veterinary Medical Experiment Station.
REFERENCES 1. Saiki,R.K., Gelfand,D.H., Stoffel,S., Scharf,S.J., Higuchi,R., Horn,G.T., Mullis,K.B. and Ehrlich,H.A. (1988) Science 239, 487-491. 2. Shirai,T., Yamaguchi,H., Ito,H., Todd,C.W. and Wallace,R.B. (1985) Nature 313, 803-806. 3. McGraw,R.A., Steffe,E.K. and Hutchinson,C.A. (1988) DNA and Protein Engineering Techniques 1, 65-67.
-
S
M
M
I
R
D
E
V
L
A
E
GAGCTGGCCG
E
A
L
P KK
AGGAGGCACT CCCCAAGAAG
101
L H F G C L GCCTCTTCTC CTTCCTCCTC GTCGCAGGAG CCACCACACT CTTCTGCCTG CTGCACTTTG
201 301 401 501 601 701
AGACTTGGCT AAACGTGGAG ATGAATGAGC TGCGAGACAC AAAGTGCTGT GGTAACTTGT
801 901
1001 1101
1201 1301
F
S
F
L
V
A
G
T
A
T
L
F
G
P
L
C
L
S
TTGTGCCTCA
100
G TAAGTGTCTC
800
CAAAGCTTCT CTCCTGGTTC TGGGCTTGGG CGAGGGTGGA GGTTAGTCCT GGGATAGCAG CAGAGGGGGA AATTCAGAGT TTGGGTTTGA GGGGAGGAGG AATGGAAGTC AAAGTGGGAT ATTTTCCAGG AAGCGTAAGG GTCTCACCTT TTTCTTTCCT CTTTCCTCCT CAG P S D K S R T S S
900
CATTCTCCTA AAAGACGGGA ACACACATAG AATGATGTGG TGAATGAATG TCTGATGCTA
CCCAGAGAGA GGCAGAAAGA ACAGACAGAC GCGATATGAG AATGGATGAA AACCTCCTCC
A P V A H V V AGCCAGTAGC CCATGTAGTA G GTAAGACTT GAAGGCTGAG CCCAGGCCTT GGAGATGATA CTCATGGCCA CATGAGACGT GGGAGGACAG TAACAGGGTC ACACTGACTT ACCCCTCCCT L
L
A
GCCCTCCTGG C P S
1501
GATGTCCTTC Q R E CCAGAGGGAG E I N L GAGATCAATC
1701
L
G
GCAGGGGGCC
200 300 400 500 600 700
1401
1601
L
G R C Q G S CCCAGGGCTC CGGAAGGTGC E E P Q R V I G GAGTGATCGG CCCCCAGAGG GAAGAG GTGA TGGGGGAAGG AGGTGTGCTG ATGGGGATGG GGAGCACACC AAAGGGTCTC TCTAAGCACT GGAGAGATGT GGGGTGTGGG TTGAGAGATG AAGGAAAGAC AGACAGATGT CTGTCACGAG GTATGAGCTG CATAAACAAC CTAGCTGTTT
A
GTGAGAGAGA TCTTGAATGA GGAATAAAGA GAAGACAGTG CTCGGTTAGG
A
Figure 1. Nucleotide derived sequences
*
S
E
T
ATGACACTG AAGCATGA fCGGGACGTG
X54000
ACKNOWLEDGEMENT
We report genomic DNA sequence of the feline tumor necrosis factor alpha. The gene segment was obtained by PCR (1) from primers: the using DNA feline genomic 5'-ATGAGCACTGAAAGCATGATC-3' (sense) and 5 '-TCACAGGGCAATGATCCCAAAGTA-3' (antisense). Primers were based on published human sequence (2) at the extremes of known coding regions. Amplification was performed in a home made thermal cycling apparatus (3). A PCR product of 1.8 kb was excised and purified from a .7% agarose gel, 5-phosphorylated with T4 polynucleotide kinase, end-repaired with Klenow fragment, cloned into the SmaI site of pUC 19, and further subcloned into M13 vectors for conventional dideoxy sequencing in both strands.
1
no.
To whom
are
N G V CCAATGGCGT T H V CACACATGTG T P E G ACCCCAGAGG P A Y TGCCTGCCTA
sequence
AATGGGAGGG GATGTGGAGA AGAGAGACAG AGAGATAAGG TCAATCGATG TTCTCCCCAA
(1772bp) of the
GTGCCACACC
T L R P L P Q L I N G L Q ATGGCCTGCA ACTAATCAAC CCTCTGCCCC AGACACTCA
1000
CTGAGGCTGT AGCAGGAGGA TGGCGAAAGG GTGGGACATA GAGGGCTAGG ATTTGGGGGC TGAGCAGGCT TGAGAAGGAC TCACTGAGCT CAAGGAGAGA GTGAAGGAAG AGCACAGGCC TTGAGGGGGA CACTCAGAAC ACAGAGGGGA CAGGAACCGG ATGTGGGGGG GGGGGGCAGA ACTTGAGGGC CAGGATGTGG AGAGTGGAAC
1100 1200 1300
CCTTCTCTCC TTCCCTCCAG
gene
correspondence should be addressed
AGAAATGGGG GAGAAGGGAC AGAGATGTGT AGAAAGAGAT GAGAACCAGA
GATCATC TTCTCGAACT CCGAGTGACA
D N Q L E L T GGAGCTGACA GACAACCAAC A I S L L T H CTCCTCACCC ACGCCATCAG K P W A E A GGGCTGAGGC CAAGCCCTGG A E S G L D F TCTTGACTTT GCCGAGTCTG
underlined.
CAGGAGGGCA GCGATGAAGA AGAGAGAGAG AAAGAGGTGA AACGGAGAGA CAG L P H CTCCCAC
K V P TGAAGGTGCC R F A CCGCTTTGCC Y E P I TATGAGCCCA Q V Y
R A N L S R Q L Q R A E G N P E CAAACCCCGA AGCTGAAGGG CAACTCCAGC GGCTGAGCCG GCGTGCCAAT T G Q G V L F Y S Q S D G L Y L I
1400
TCTACTCCCA GGTCCTCTTC ACGGGCCAAG S P C S A I K N L L CAACCTCCTC TCTGCCATCA AGAGCCCTTG L S T G D R Q L E K CAACTGGAGA AGGGTGATCG ACTCAGCACT
1600
CTCAGACGGG V S Y Q GTTTCCTACC Y L G TCTACCTGGG F G I
GGCAAGTCIA C
GGGATC
CTGTACCTCA T K V AGACCAAGGT G V F AGGGGTCTTC I A L *
A
T1 G&
1500
1700 1772
for feline tumor necrosis factor alpha. Predicted translation is shown above the nucleotide sequence. Primer
.. 1990 Oxford University Press
of feline tumor necrosis factor alpha
Gene sequence
Royal A.McGraw*, Bradford W.Coffee, Cynthia M.Ottol, Roman T.Drews and Clarence A.Rawlings1 Departments of Physiology and Pharmacology and 'Small Animal Medicine, College of Veterinary Medicine, University of Georgia, Athens, GA 30602, USA EMBL accession
Submitted August 3, 1990
This work was supported by the University of Georgia Veterinary Medical Experiment Station.
REFERENCES 1. Saiki,R.K., Gelfand,D.H., Stoffel,S., Scharf,S.J., Higuchi,R., Horn,G.T., Mullis,K.B. and Ehrlich,H.A. (1988) Science 239, 487-491. 2. Shirai,T., Yamaguchi,H., Ito,H., Todd,C.W. and Wallace,R.B. (1985) Nature 313, 803-806. 3. McGraw,R.A., Steffe,E.K. and Hutchinson,C.A. (1988) DNA and Protein Engineering Techniques 1, 65-67.
-
S
M
M
I
R
D
E
V
L
A
E
GAGCTGGCCG
E
A
L
P KK
AGGAGGCACT CCCCAAGAAG
101
L H F G C L GCCTCTTCTC CTTCCTCCTC GTCGCAGGAG CCACCACACT CTTCTGCCTG CTGCACTTTG
201 301 401 501 601 701
AGACTTGGCT AAACGTGGAG ATGAATGAGC TGCGAGACAC AAAGTGCTGT GGTAACTTGT
801 901
1001 1101
1201 1301
F
S
F
L
V
A
G
T
A
T
L
F
G
P
L
C
L
S
TTGTGCCTCA
100
G TAAGTGTCTC
800
CAAAGCTTCT CTCCTGGTTC TGGGCTTGGG CGAGGGTGGA GGTTAGTCCT GGGATAGCAG CAGAGGGGGA AATTCAGAGT TTGGGTTTGA GGGGAGGAGG AATGGAAGTC AAAGTGGGAT ATTTTCCAGG AAGCGTAAGG GTCTCACCTT TTTCTTTCCT CTTTCCTCCT CAG P S D K S R T S S
900
CATTCTCCTA AAAGACGGGA ACACACATAG AATGATGTGG TGAATGAATG TCTGATGCTA
CCCAGAGAGA GGCAGAAAGA ACAGACAGAC GCGATATGAG AATGGATGAA AACCTCCTCC
A P V A H V V AGCCAGTAGC CCATGTAGTA G GTAAGACTT GAAGGCTGAG CCCAGGCCTT GGAGATGATA CTCATGGCCA CATGAGACGT GGGAGGACAG TAACAGGGTC ACACTGACTT ACCCCTCCCT L
L
A
GCCCTCCTGG C P S
1501
GATGTCCTTC Q R E CCAGAGGGAG E I N L GAGATCAATC
1701
L
G
GCAGGGGGCC
200 300 400 500 600 700
1401
1601
L
G R C Q G S CCCAGGGCTC CGGAAGGTGC E E P Q R V I G GAGTGATCGG CCCCCAGAGG GAAGAG GTGA TGGGGGAAGG AGGTGTGCTG ATGGGGATGG GGAGCACACC AAAGGGTCTC TCTAAGCACT GGAGAGATGT GGGGTGTGGG TTGAGAGATG AAGGAAAGAC AGACAGATGT CTGTCACGAG GTATGAGCTG CATAAACAAC CTAGCTGTTT
A
GTGAGAGAGA TCTTGAATGA GGAATAAAGA GAAGACAGTG CTCGGTTAGG
A
Figure 1. Nucleotide derived sequences
*
S
E
T
ATGACACTG AAGCATGA fCGGGACGTG
X54000
ACKNOWLEDGEMENT
We report genomic DNA sequence of the feline tumor necrosis factor alpha. The gene segment was obtained by PCR (1) from primers: the using DNA feline genomic 5'-ATGAGCACTGAAAGCATGATC-3' (sense) and 5 '-TCACAGGGCAATGATCCCAAAGTA-3' (antisense). Primers were based on published human sequence (2) at the extremes of known coding regions. Amplification was performed in a home made thermal cycling apparatus (3). A PCR product of 1.8 kb was excised and purified from a .7% agarose gel, 5-phosphorylated with T4 polynucleotide kinase, end-repaired with Klenow fragment, cloned into the SmaI site of pUC 19, and further subcloned into M13 vectors for conventional dideoxy sequencing in both strands.
1
no.
To whom
are
N G V CCAATGGCGT T H V CACACATGTG T P E G ACCCCAGAGG P A Y TGCCTGCCTA
sequence
AATGGGAGGG GATGTGGAGA AGAGAGACAG AGAGATAAGG TCAATCGATG TTCTCCCCAA
(1772bp) of the
GTGCCACACC
T L R P L P Q L I N G L Q ATGGCCTGCA ACTAATCAAC CCTCTGCCCC AGACACTCA
1000
CTGAGGCTGT AGCAGGAGGA TGGCGAAAGG GTGGGACATA GAGGGCTAGG ATTTGGGGGC TGAGCAGGCT TGAGAAGGAC TCACTGAGCT CAAGGAGAGA GTGAAGGAAG AGCACAGGCC TTGAGGGGGA CACTCAGAAC ACAGAGGGGA CAGGAACCGG ATGTGGGGGG GGGGGGCAGA ACTTGAGGGC CAGGATGTGG AGAGTGGAAC
1100 1200 1300
CCTTCTCTCC TTCCCTCCAG
gene
correspondence should be addressed
AGAAATGGGG GAGAAGGGAC AGAGATGTGT AGAAAGAGAT GAGAACCAGA
GATCATC TTCTCGAACT CCGAGTGACA
D N Q L E L T GGAGCTGACA GACAACCAAC A I S L L T H CTCCTCACCC ACGCCATCAG K P W A E A GGGCTGAGGC CAAGCCCTGG A E S G L D F TCTTGACTTT GCCGAGTCTG
underlined.
CAGGAGGGCA GCGATGAAGA AGAGAGAGAG AAAGAGGTGA AACGGAGAGA CAG L P H CTCCCAC
K V P TGAAGGTGCC R F A CCGCTTTGCC Y E P I TATGAGCCCA Q V Y
R A N L S R Q L Q R A E G N P E CAAACCCCGA AGCTGAAGGG CAACTCCAGC GGCTGAGCCG GCGTGCCAAT T G Q G V L F Y S Q S D G L Y L I
1400
TCTACTCCCA GGTCCTCTTC ACGGGCCAAG S P C S A I K N L L CAACCTCCTC TCTGCCATCA AGAGCCCTTG L S T G D R Q L E K CAACTGGAGA AGGGTGATCG ACTCAGCACT
1600
CTCAGACGGG V S Y Q GTTTCCTACC Y L G TCTACCTGGG F G I
GGCAAGTCIA C
GGGATC
CTGTACCTCA T K V AGACCAAGGT G V F AGGGGTCTTC I A L *
A
T1 G&
1500
1700 1772
for feline tumor necrosis factor alpha. Predicted translation is shown above the nucleotide sequence. Primer
