Download Bz gene identification

Worksheet: Identify the bz gene in DNA

Assuming the bz gene could be a simple ORF gene try to identify it by detecting and
analyzing the ORFs in the sequence.
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Go to http://www.bioservers.orgFind SEQUENCE SERVER ENTER
Click MANAGE GROUPS
Find Sequence sources, click Classes, then Public.
Find Jumping Genes Across Kingdoms
Check the box to the left, click OK
Click the title for the first entry and set it to corn, purple endosperm; wt
Click View
Highlight and copy the entire sequence.
Open http://www.dnai.org/geneboy
In the Sequences panel click Your Sequence
Paste the sequence into the central window. Replace the header Your Sequence
with a name of your choosing (i.e. corn bz gene region). Click Save Sequence
How long is the sequence? __2221_ bp
In the Operations panel click Find Genes, then ORFs
Click Reverse.
Record in the table below the ORFs indicated by Gene Boy.
ORF
ORF 1
ORF 2
ORF 3
ORF 4
ORF 5

RF
+1 _
+2 _
-1 _
-1 _
-3 _
From – To
247 - 834
842 - 1762
220 - 819
1117 - 1500
890 - 1867
_
_
_
_
_
Length [bp]
588
_
921
_
600
_
384
_
978
_
Protein length [aa]
195
_
306
_
299
_
127
_
325
_
The protein sequencing lab provides you with the amino acid for the protein product of
the bz-gene (see Attachment 1).
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How many amino acids long is it? ___ 471 aa ____________
How many nucleotides would be required to encode a protein of this length? 1413
Could it be encoded by any of the ORFs determined above? Nope
What do you think might be going on? At what point may we have made a wrong
assumption?
_________________________________________________________________
_ Assuming the gene is a single ORF gene. Instead, it may be a spliced gene. _
_________________________________________________________________
_________________________________________________________________

Using the DNA sequence from Sequence Server and the translation tool at
http://www.dnalc.org/bioinformatics/2003/2003_dnalc_nucleotide_analyzer.htm#translat
or, the Bioinformatics Department has provided you with a translation of the sequence in
all three forward reading frames (see Attachment 2). Detect the amino acid sequence for
the bz protein product (Attachment 1) in these three deduced amino acid sequences.
Highlight in the translated sequences the amino acid stretches that are entailed in the BZ
protein sequence.

In order to identify the bz gene in the DNA sequence highlight the nucleotide stretches
that correspond to the highlighted amino acid stretches. If necessary consult the genetic
code table in Attachment 3.

Discuss the structure of the gene:
o What is the structure of the bz gene?
__It consists of two exons and one intron
________________________
o At what position are the start and stop codons located?
___Start:____247 – 249 ________
Stop:___1760 – 1762
__________
o How many substructures does the coding region of the gene consist of?
How long are these substructures? Are they divisible by three?
____Two exons;_CDS 1: 523 bp, CDS 2: 890 bp ________________________
o Concatenate the coding substructures. How long is this sequence? Is it a multiple
of three? Would it be able to encode a protein of the length of the BZ protein?
_____Total length CDS: 1413; encodes 471 amino acids ___________________

Use the Internet sites at http://wwwmgs.bionet.nsc.ru/mgs/programs/bdna/tata_bdna.html
and http://rulai.cshl.org/tools/polyadq/polyadq_form.html for the prediction of TATAboxes and PolyA Signal, respectively.
______ see annotation in the sequence below ___________________________
_________________________________________________________________
_________________________________________________________________

Finally, run the sequence through the two gene prediction programs listed in Gene Boy
under WWW Tools  Gene Prediction.
_________________________________________________________________
_________________________________________________________________
_________________________________________________________________

Discuss the results by comparing them with the annotation for the gene at.
http://www.ncbi.nlm.nih.gov/entrez/viewer.fcgi?db=nucleotide&val=22361
_________________________________________________________________
_________________________________________________________________
Attachment 1: Zea mays bz gene product; 471 amino acids
---------+---------+---------+---------+---------+---------+
MAPADGESSPPPHVAVVAFPFSSHAAVLLSIARALAAAAAPSGATLSFLSTASSLAQLRK 60
---------+---------+---------+---------+---------+---------+
ASSASAGHGLPGNLRFVEVPDGAPAAEETVPVPRQMQLFMEAAEAGGVKAWLEAARAAAG 120
---------+---------+---------+---------+---------+---------+
GARVTCVVGDAFVWPAADAAASAGAPWVPVWTAASCALLAHIRTDALREDVGDQAANRVD 180
---------+---------+---------+---------+---------+---------+
GLLISHPGLASYRVRDLPDGVVSGDFNYVINLLVHRMGQCLPRSAAAVALNTFPGLDPPD 240
---------+---------+---------+---------+---------+---------+
VTAALAEILPNCVPFGPYHLLLAEDDADTAAPADPHGCLAWLGRQPARGVAYVSFGTVAC 300
---------+---------+---------+---------+---------+---------+
PRPDELRELAAGLEDSGAPFLWSLREDSWPHLPPGFLDRAAGTGSGLVVPWAPQVAVLRH 360
---------+---------+---------+---------+---------+---------+
PSVGAFVTHAGWASVLEGLSSGVPMACRPFFGDQRMNARSVAHVWGFGAAFEGAMTSAGV 420
---------+---------+---------+---------+---------+ATAVEELLRGEEGARMRARAKELQALVAEAFGPGGECRKNFDRFVEIVCRA 471
Attachment 2: bz gene, Zea mays, 2221 nucleotides
1--------+---------+---------+---------+---------+---------+---------+---------+---------+---------+-DNA: GGTCCCCAAACTCCACGGCACCAACAGCTAAGCCCGATGCGCTGCGTGCGCGGCGATCCAACCGCCGGCTCACCTAAAAATTTCGGCACGTCTAACTGCGAC
+1: G P Q T P R H Q Q L S P M R C V R G D P T A G S P K N F G T S N C D
+2: V P K L H G T N S * A R C A A C A A I Q P P A H L K I S A R L T A T
+3:
S P N S T A P T A K P D A L R A R R S N R R L T * K F R H V * L R L
102
------------------------------------------------------------------------------------------------------------------------------103----+---------+---------+---------+---------+---------+---------+---------+---------+---------+---DNA: TGGCAGGTGCGCACGCGTGGTCGCGCGGAATAAAGCGGACACGTTGCGCCCCCAGCGAAGCCCGCACGCATCGCATTCGCATCGCATCGCAGGTCGCATCCG
+1: W Q V R T R G R A E * S G H V A P P A K P A R I A F A S H R R S H P
+2: G R C A R V V A R N K A D T L R P Q R S P H A S H S H R I A G R I R
+3:
A G A H A W S R G I K R T R C A P S E A R T H R I R I A S Q V A S D
204
------------------------------------------------------------------------------------------------------------------------------205--+---------+---------+---------+---------+---------+---------+---------+---------+---------+-----DNA: ACGCTAGCGGCTAGCCTAGCCGAACAGCCTGAGCGCGCGAAGATGGCGCCCGCCGACGGCGAGTCCTCCCCGCCGCCGCACGTGGCCGTGGTCGCCTTCCCG
+1: T L A A S L A E Q P E R A K M A P A D G E S S P P P H V A V V A F P
+2: R * R L A * P N S L S A R R W R P P T A S P P R R R T W P W S P S R
+3:
A S G * P S R T A * A R E D G A R R R R V L P A A A R G R G R L P V
306
------------------------------------------------------------------------------------------------------------------------------3--+---------+---------+---------+---------+---------+---------+---------+---------+---------+-------DNA: TTCAGCTCCCACGCGGCGGTGCTGCTCTCCATCGCGCGCGCCCTGGCTGCCGCCGCGGCGCCGTCCGGGGCCACGCTCTCGTTCCTCTCCACCGCGTCCTCC
+1: F S S H A A V L L S I A R A L A A A A A P S G A T L S F L S T A S S
+2: S A P T R R C C S P S R A P W L P P R R R P G P R S R S S P P R P P
+3:
Q L P R G G A A L H R A R P G C R R G A V R G H A L V P L H R V L P
408
------------------------------------------------------------------------------------------------------------------------------409--------+---------+---------+---------+---------+---------+---------+---------+---------+---------+
DNA: CTCGCGCAGCTCCGCAAGGCCAGCAGCGCCTCCGCCGGGCACGGGCTCCCGGGGAACCTGCGCTTCGTCGAGGTACCGGACGGCGCGCCCGCGGCCGAGGAG
+1: L A Q L R K A S S A S A G H G L P G N L R F V E V P D G A P A A E E
+2: S R S S A R P A A P P P G T G S R G T C A S S R Y R T A R P R P R R
+3:
R A A P Q G Q Q R L R R A R A P G E P A L R R G T G R R A R G R G D
510
------------------------------------------------------------------------------------------------------------------------------511------+---------+---------+---------+---------+---------+---------+---------+---------+---------+-DNA: ACCGTGCCGGTGCCGCGGCAGATGCAGCTGTTCATGGAGGCCGCGGAGGCCGGCGGGGTGAAGGCCTGGCTGGAGGCGGCCCGCGCCGCGGCGGGCGGCGCC
+1: T V P V P R Q M Q L F M E A A E A G G V K A W L E A A R A A A G G A
+2: P C R C R G R C S C S W R P R R P A G * R P G W R R P A P R R A A P
+3:
R A G A A A D A A V H G G R G G R R G E G L A G G G P R R G G R R Q
612
613----+---------+---------+---------+---------+---------+---------+---------+---------+---------+---DNA: AGGGTGACCTGCGTGGTGGGCGACGCGTTCGTGTGGCCGGCGGCGGACGCGGCCGCCTCCGCGGGGGCGCCGTGGGTGCCGGTGTGGACGGCCGCGTCGTGC
+1: R V T C V V G D A F V W P A A D A A A S A G A P W V P V W T A A S C
+2: G * P A W W A T R S C G R R R T R P P P R G R R G C R C G R P R R A
+3:
G D L R G G R R V R V A G G G R G R L R G G A V G A G V D G R V V R
714
------------------------------------------------------------------------------------------------------------------------------715--+---------+---------+---------+---------+---------+---------+---------+---------+---------+-----DNA: GCGCTCCTGGCGCACATCCGCACCGACGCGCTCCGGGAGGACGTTGGCGACCAGGGTGCGTTGGATTCTACTACTACTACTTCTCTCCCTTCCTTGTCCCTT
+1: A L L A H I R T D A L R E D V G D Q G A L D S T T T T S L P S L S L
+2: R S W R T S A P T R S G R T L A T R V R W I L L L L L L S L P C P F
+3:
A P G A H P H R R A P G G R W R P G C V G F Y Y Y Y F S P F L V P S
816
------------------------------------------------------------------------------------------------------------------------------817+---------+---------+---------+---------+---------+---------+---------+---------+---------+-------DNA: CATTGCGCGCGGGTTTGATGATCGAATGGCTGTTGCATTTCCATCGTTCGCAGCAGCAAACAGGGTGGACGGGCTACTGATCTCCCACCCGGGCCTCGCCAG
+1: H C A R V * * S N G C C I S I V R S S K Q G G R A T D L P P G P R Q
+2: I A R G F D D R M A V A F P S F A A A N R V D G L L I S H P G L A S
+3:
L R A G L M I E W L L H F H R S Q Q Q T G W T G Y * S P T R A S P A
918
------------------------------------------------------------------------------------------------------------------------------919--------+---------+---------+---------+---------+---------+---------+---------+---------+---------+
DNA: CTACCGCGTCCGTGACCTCCCAGACGGCGTCGTCTCCGGCGACTTCAACTACGTCATCAACCTCCTCGTCCACCGCATGGGGCAGTGCCTCCCGCGCTCTGC
+1: L P R P * P P R R R R L R R L Q L R H Q P P R P P H G A V P P A L C
+2: Y R V R D L P D G V V S G D F N Y V I N L L V H R M G Q C L P R S A
+3:
T A S V T S Q T A S S P A T S T T S S T S S S T A W G S A S R A L P
1020
------------------------------------------------------------------------------------------------------------------------------1021-----+---------+---------+---------+---------+---------+---------+---------+---------+---------+-DNA: CGCCGCCGTGGCACTCAACACGTTCCCAGGCCTGGACCCGCCCGACGTCACCGCGGCGCTCGCGGAGATCCTGCCCAACTGCGTCCCGTTCGGCCCCTACCA
+1: R R R G T Q H V P R P G P A R R H R G A R G D P A Q L R P V R P L P
+2: A A V A L N T F P G L D P P D V T A A L A E I L P N C V P F G P Y H
+3:
P P W H S T R S Q A W T R P T S P R R S R R S C P T A S R S A P T T
1122
------------------------------------------------------------------------------------------------------------------------------1123---+---------+---------+---------+---------+---------+---------+---------+---------+---------+---DNA: CCTCCTCCTCGCCGAGGACGACGCCGACACCGCCGCACCAGCCGACCCGCACGGCTGCCTCGCCTGGCTGGGCCGCCAACCCGCGCGCGGCGTCGCGTACGT
+1: P P P R R G R R R H R R T S R P A R L P R L A G P P T R A R R R V R
+2: L L L A E D D A D T A A P A D P H G C L A W L G R Q P A R G V A Y V
+3:
S S S P R T T P T P P H Q P T R T A A S P G W A A N P R A A S R T S
1224
1225-+---------+---------+---------+---------+---------+---------+---------+---------+---------+-----DNA: CAGCTTCGGCACGGTGGCGTGCCCGCGGCCCGACGAGCTCCGCGAGCTGGCGGCCGGGCTGGAGGACTCGGGCGCGCCGTTCCTGTGGTCGCTGCGCGAGGA
+1: Q L R H G G V P A A R R A P R A G G R A G G L G R A V P V V A A R G
+2: S F G T V A C P R P D E L R E L A A G L E D S G A P F L W S L R E D
+3:
A S A R W R A R G P T S S A S W R P G W R T R A R R S C G R C A R T
1326
------------------------------------------------------------------------------------------------------------------------------1327---------+---------+---------+---------+---------+---------+---------+---------+---------+-------DNA: CTCGTGGCCGCACCTCCCGCCGGGTTTCCTGGACCGCGCCGCGGGCACCGGGTCCGGGCTCGTGGTGCCCTGGGCGCCGCAGGTGGCCGTGCTGCGCCACCC
+1: L V A A P P A G F P G P R R G H R V R A R G A L G A A G G R A A P P
+2: S W P H L P P G F L D R A A G T G S G L V V P W A P Q V A V L R H P
+3:
R G R T S R R V S W T A P R A P G P G S W C P G R R R W P C C A T L
1428
------------------------------------------------------------------------------------------------------------------------------1429-------+---------+---------+---------+---------+---------+---------+---------+---------+---------+
DNA: TTCCGTGGGCGCGTTCGTGACGCACGCCGGGTGGGCGTCGGTGCTGGAGGGCTTGTCCAGCGGGGTGCCCATGGCGTGCCGCCCCTTCTTCGGCGACCAGCG
+1: F R G R V R D A R R V G V G A G G L V Q R G A H G V P P L L R R P A
+2: S V G A F V T H A G W A S V L E G L S S G V P M A C R P F F G D Q R
+3:
P W A R S * R T P G G R R C W R A C P A G C P W R A A P S S A T S G
1530
------------------------------------------------------------------------------------------------------------------------------1531-----+---------+---------+---------+---------+---------+---------+---------+---------+---------+-DNA: GATGAACGCGCGGTCCGTGGCGCACGTGTGGGGGTTCGGCGCCGCGTTCGAGGGCGCTATGACGAGCGCCGGAGTGGCCACGGCCGTGGAGGAGCTGCTGCG
+1: D E R A V R G A R V G V R R R V R G R Y D E R R S G H G R G G A A A
+2: M N A R S V A H V W G F G A A F E G A M T S A G V A T A V E E L L R
+3:
* T R G P W R T C G G S A P R S R A L * R A P E W P R P W R S C C A
1632
------------------------------------------------------------------------------------------------------------------------------1633---+---------+---------+---------+---------+---------+---------+---------+---------+---------+---DNA: CGGGGAGGAAGGGGCGCGGATGAGGGCAAGGGCCAAGGAGCTGCAGGCCTTGGTGGCCGAGGCGTTCGGGCCAGGCGGTGAGTGCAGGAAGAACTTCGACAG
+1: R G G R G A D E G K G Q G A A G L G G R G V R A R R * V Q E E L R Q
+2: G E E G A R M R A R A K E L Q A L V A E A F G P G G E C R K N F D R
+3:
G R K G R G * G Q G P R S C R P W W P R R S G Q A V S A G R T S T G
1734
------------------------------------------------------------------------------------------------------------------------------1735-+---------+---------+---------+---------+---------+---------+---------+---------+---------+-----DNA: GTTCGTCGAGATAGTCTGTCGCGCGTGAAAGGTCGTCTTGCTGTTCAGAGGTTTTACCAACAGAAGAACATAATGAATTGGATGGCATGCTACGTCGTATTC
+1: V R R D S L S R V K G R L A V Q R F Y Q Q K N I M N W M A C Y V V F
+2: F V E I V C R A * K V V L L F R G F T N R R T * * I G W H A T S Y S
+3:
S S R * S V A R E R S S C C S E V L P T E E H N E L D G M L R R I L
1836
1837---------+---------+---------+---------+---------+---------+---------+---------+---------+-------DNA: TCTTTTTTTGTTGATCCCTGAGTTGATACATTTTGTACTTGATACATGAGTTGCAGCAGCAGCAGCAACAGCCTTCTGTACCTTGGCTTTGGATCTGTATTC
+1: S F F V D P * V D T F C T * Y M S C S S S S N S L L Y L G F G S V F
+2: L F L L I P E L I H F V L D T * V A A A A A T A F C T L A L D L Y S
+3:
F F C * S L S * Y I L Y L I H E L Q Q Q Q Q Q P S V P W L W I C I L
1938
------------------------------------------------------------------------------------------------------------------------------1939-------+---------+---------+---------+---------+---------+---------+---------+---------+---------+
DNA: TTGTCACCAGTTATCTGAAAGCATCAATAACCTTCTGTCTTCTAGCAGTTGCCTCTCCAGATTGCCAAAATAGCATTTATTATAAGGTCTTATGCAATGTTT
+1: L S P V I * K H Q * P S V F * Q L P L Q I A K I A F I I R S Y A M F
+2: C H Q L S E S I N N L L S S S S C L S R L P K * H L L * G L M Q C F
+3:
V T S Y L K A S I T F C L L A V A S P D C Q N S I Y Y K V L C N V F
2040
------------------------------------------------------------------------------------------------------------------------------2041-----+---------+---------+---------+---------+---------+---------+---------+---------+---------+-DNA: TCAGATTGTTCCGATTAAATCTACGATTAGCATTTTAGCCCAGCAGTCCAGCCCATTGAAGGCTTATTCAGTTATTTTTAATCCATATAAATCAAAAAAGAT
+1: S D C S D * I Y D * H F S P A V Q P I E G L F S Y F * S I * I K K D
+2: Q I V P I K S T I S I L A Q Q S S P L K A Y S V I F N P Y K S K K I
+3:
R L F R L N L R L A F * P S S P A H * R L I Q L F L I H I N Q K R L
2142
------------------------------------------------------------------------------------------------------------------------------2143---+---------+---------+---------+---------+---------+---------+---------+DNA: TGATATAGATTAGAAAATATTTTAGTTTACTAGGAATTAAAACCCCTCAATTTTTCTTAATCCATATAAATTGTGGCAG
+1: * Y R L E N I L V Y * E L K P L N F S * S I * I V A
+2: D I D * K I F * F T R N * N P S I F L N P Y K L W Q
+3:
I * I R K Y F S L L G I K T P Q F F L I H I N C G
2221
-------------------------------------------------------------------------------------------------------------------------------
Attachment 3: Genetic Code (http://psyche.uthct.edu/shaun/SBlack/geneticd.html)
Second Position of Codon
T
C
A
G
TTT Phe [F] TCT Ser [S]
TTC Phe [F] TCC Ser [S]
T
TTA Leu [L] TCA Ser [S]
F
TTG Leu [L] TCG Ser [S]
i
r
CTT Leu [L] CCT Pro [P]
s
CTC Leu [L] CCC Pro [P]
t C
CTA Leu [L] CCA Pro [P]
CTG Leu [L] CCG Pro [P]
P
TAT Tyr [Y]
TAC Tyr [Y]
TAA Ter [end]
TAG Ter [end]
TGT Cys [C]
TGC Cys [C]
TGA Ter [end]
TGG Trp [W]
CAT His [H]
CAC His [H]
CAA Gln [Q]
CAG Gln [Q]
CGT Arg [R]
CGC Arg [R]
CGA Arg [R]
CGG Arg [R]
T
C
A
G T
h
T i
C r
d
A
G P
o
ATT Ile [I]
s
ATC Ile [I]
i A
ATA Ile [I]
t
ATG Met [M]
i
o
GTT Val [V]
n
GTC Val [V]
G
GTA Val [V]
GTG Val [V]
ACT Thr [T]
ACC Thr [T]
ACA Thr [T]
ACG Thr [T]
AAT Asn [N]
AAC Asn [N]
AAA Lys [K]
AAG Lys [K]
AGT Ser [S]
AGC Ser [S]
AGA Arg [R]
AGG Arg [R]
T
C
A
G
GCT Ala [A]
GCC Ala [A]
GCA Ala [A]
GCG Ala [A]
GAT Asp [D]
GAC Asp [D]
GAA Glu [E]
GAG Glu [E]
GGT Gly [G]
GGC Gly [G]
GGA Gly [G]
GGG Gly [G]
o
s
i
t
i
o
T n
C
A
G
An explanation of the Genetic Code: DNA is a two-stranded molecule. Each strand is a
polynucleotide composed of A (adenosine), T (thymidine), C (cytidine), and G (guanosine) residues
polymerized by "dehydration" synthesis in linear chains with specific sequences. Each strand has
polarity, such that the 5'-hydroxyl (or 5'-phospho) group of the first nucleotide begins the strand and
the 3'-hydroxyl group of the final nucleotide ends the strand; accordingly, we say that this strand
runs 5' to 3' ("Five prime to three prime") . It is also essential to know that the two strands of DNA
run antiparallel such that one strand runs 5' -> 3' while the other one runs 3' -> 5'. At each nucleotide
residue along the double-stranded DNA molecule, the nucleotides are complementary. That is, A
forms two hydrogen-bonds with T; C forms three hydrogen bonds with G. In most cases the twostranded, antiparallel, complementary DNA molecule folds to form a helical structure which
resembles a spiral staircase. This is the reason why DNA has been referred to as the "Double
Helix".
One strand of DNA holds the information that codes for various genes; this strand is often called the
template strand or antisense strand (containing anticodons). The other, and complementary, strand
is called the coding strand or sense strand (containing codons). Since mRNA is made from the
template strand, it has the same information as the coding strand. The table above refers to triplet
nucleotide codons along the sequence of the coding or sense strand of DNA as it runs 5' -> 3'; the
code for the mRNA would be identical but for the fact that RNA contains U (uridine) rather than T.
An example of two complementary strands of DNA would be:
(5' -> 3') ATGGAATTCTCGCTC
(Coding, sense strand)
(3' <- 5') TACCTTAAGAGCGAG
(Template, antisense strand)
(5' -> 3') AUGGAAUUCUCGCUC
(mRNA made from Template strand)
Since amino acid residues of proteins are specified as triplet codons, the protein sequence made
from the above example would be Met-Glu-Phe-Ser-Leu... (MEFSL...).
Practically, codons are "decoded" by transfer RNAs (tRNA) which interact with a ribosome-bound
messenger RNA (mRNA) containing the coding sequence. There are 64 different tRNAs, each of
which has an anticodon loop (used to recognize codons in the mRNA). 61 of these have a bound
amino acyl residue; the appropriate "charged" tRNA binds to the respective next codon in the mRNA
and the ribosome catalyzes the transfer of the amino acid from the tRNA to the growing (nascent)
protein/polypeptide chain. The remaining 3 codons are used for "punctuation"; that is, they signal
the termination (the end) of the growing polypeptide chain.
Lastly, the Genetic Code in the table above has also been called "The Universal Genetic Code". It is
known as "universal", because it is used by all known organisms as a code for DNA, mRNA, and
tRNA. The universality of the genetic code encompases animals (including humans), plants, fungi,
archaea, bacteria, and viruses. However, all rules have their exceptions, and such is the case with
the Genetic Code; small variations in the code exist in mitochondria and certain microbes.
Nonetheless, it should be emphasized that these variances represent only a small fraction of known
cases, and that the Genetic Code applies quite broadly, certainly to all known nuclear genes.
Suggestions for additions or changes can be sent to Dr. Shaun D. Black
Last update August 25, 1998
Shaun D. Black, University of Texas Health Center at Tyler