7831

• UCREFRP Catalog Number: 7831
• Author: Dowling, T. E., et al.
• Title: Mitochondrial DNA Variability in the Endangered Razorback Sucker ‎(
• USFW Year: 1996
• Copyright Material: YES

122

mtDNA Diversity in Razorback Sucker

tive to its source would indicate a change in genetic com-
position with potentially deleterious effects, especially in
. a species already subject to other environmental stresses.

Materials and Methods

Description of Source and Hatchery Populations

A broodstock was obtained in 1981-1982 from Lake Mo-
have, a mainstem impoundment on the Colorado River
created in the early 1950s by closure of Davis Dam. The
reservoir consists of an upstream riverine reach rv60 km
long that grades into a wider lower basin ""'48 km long
(maximum width 6.4 km). There are no direct tributar-
ies to the reservoir that contained razorback sucker popula-
tions prior to impoundment, and, based on age structure
(McCarthy & Minckley 1987; Minckley et al. 1991), re-
cruitment was negligible after an initial, in situ popula-
tion explosion. At the time of sampling (1987-1989) the
source population consisted of approximately 55,000 in-
dividuals, presumably representing progeny of residents
in this reach of river at the time of impoundment. The
population is declining; more recent estimates indicate
fewer than 23,300 adult razorback suckers remained in
1993 (Marsh 1994). Although scattered throughout, fish
are concentrated in upper parts of the lower basin. They
are vulnerable to capture when aggregated near shore
for spawning in late autumn through spring, but are not
as readily collected at other times when they are pre-
sumably dispersed throughout the basin. The species
has made up 10-30% of net catches in spawning areas
over the past 20 years (Minckley 1983; Minckley et al.
1991).
Procedures for the culture of razorback suckers are
described in Toney (1974), Inslee (1982), and Hamman
(1985). Adult fish are held in outdoor ponds and trans-
ferred to indoor holding facilities in spring for spawning.
Ovulation is induced by intramuscular injection of hu-
man chorionic gonadotropin (up to three 220 IU per kg
injections at 24-hour intervals). When necessary to in-
duce or maintain milt production, males are injected
with 660 IU common carp (Cyprinus carpio) pituitary
extract. Eggs are stripped manually, fertilized, and incu-
bated in hatchery trays. Production per female (fecun-
dity) is determined gravimetrically. Embryo viability (used
to estimate expected hatch) is assessed 48 hours after
fertilization. Actual larval production is measured by wa-
ter displacement after hatching is complete.

Isolation and Analysis of mtDNA

Genetic variation in razorback suckers was character-
ized by isolation and restriction endonuclease analysis of
mtDNA from samples derived from two sources: gamete
samples (ova or milt) from 25 individuals from Lake Mo-

Conservation Bio106'Y
Volume 10, No. I, February 1996

Dowling et ai.

have (captured in 1987-1989), and tissues (heart, liver,
gonad where available) from 31 juveniles representing
three year-classes produced at DNFH (1987 and 1990, n =
10; 1989, n = 11) and sampled in 1990. Mitochondrial
DNA was isolated by either phenol-chlorofonn extrac-
tion (Chapman & Powers 1984) or equilibrium-density
ultracentrifugation (Dowling et al. 1990). The fanner
was used initially on a small number of egg samples, and
the latter was used to purify intact, circular mtDNA from
the remaining samples.
The following restriction enzymes were used to char-
acterize mtDNAs: BanI (GGPyPuCC), BstEII (GGTNACC),
HinfI (GANTC), HinPI (GCGC), MboI (GATC), NheI
(GCTAGC), SerFI (CCNGG), and TaqI (fCAG). The re-
sulting cleavage fragments were end-labeled using all
four a32P-dNTPs, electrophoretically separated on 1.0-
1.5% agarose and 4.0% polyacrylamide gels, and visual-
ized by autoradiography (Dowling et al. 1990). Distinc-
tive restriction-fragment patterns for a specific enzyme
were identified by acronym, which may include a letter
and prime (') mark. Acronyms were assigned by order
of discovery, not pattern similarity. The composite
haplotype for each individual is identified by an eight-

Table 1. Haplotypic identification and distribution among hatchery
year classes of razorback sucker and their source population, Lake
Mohave, Arizona-Nevada.

Lake
Haplotype * Mohave 1987 1989 1990
I. AAAABA'AA 3
2. ABADA' AAA 3 1
3. BCBBCA'AA 1
4. ACAEA'A'BA 2
5. ACAAA' A'BA 1
6. ACEA'HA'AC 2 3
7. ACFABEAA 1
8. EEAABA'BA 1
9. FCAAAA'BA 1
10.ACAAAAAA 1 2 1
11. ADAADCAA 1
12. ACAAFDAA 1
13. BFA' AKA' AA 1
14. BCAFIA'AA 2
15. ACAG)A'BA 1
16. ACDCFBAB 2 3 5
17. ACAA'HA' AC 1
18. )CIAAA' AA 1 1
19. DAAABA'AA 2
20.)CIALA'AA 1
21. IGAFMA'AA I
22. ACGFAA'AA 2
23. GAAABA' AA 3
24. )CHAAA' AA 2
25. )CAAAAAA 1
26. BCAAAA'AA 1
27. BRABIA' AD 1
Total 25 10 11 10
*Tbe composite baplotype is defined by tbe following enzymes
(in order): Mbol, HinPI, HinfI, Taql, SerF!, NhcI, BstEII, Bant