76 
Fishery Bulletin 1 12(1) 
Table 1 
Locus name, primer sequences, microsatellite motif, and expected product length (in base pairs) for the 14 novel microsatel- 
lite loci developed from a Gulf Menhaden ( Brevoortia patronus) genomic DNA library. The library was constructed using two 
individuals collected with a bag seine along the shoreline near Freeport, Texas, in spring 2012. Each locus has a GenBank 
accession number that can be accessed at the National Center for Biotechnology Information (www.ncbi.nlm.nih.gov; accessed 
16 December 2012). 
Locus 
Forward primer 
Reverse primer 
Motif 
Expected 
PCR 
length 
GenBank 
accession 
number 
Bp003 
ACTGCATGGTCACTAGCACT 
C AC C ACTGC AC C GC AGC ATGCTGTC 
(CAGA)9 
170 
KC331110 
Bp017 
CATTTGGATCAAGAGTATCTATCC 
CATTCTGCCACACCTCACATAGG 
(TCTG)5 
150 
KC331112 
Bp039 
GCGTTGACGCGGAGTAATGATCCAG 
GATCTCGACTGACTTACAATGCGGC 
(GTGA)6 
220 
KC331113 
Bpl21 
ACTCAAAAGGGCCTGTCTCACGGAG 
GCTGATTATAGTCTATAGGTTGGAC 
(TCTA)19 
200 
KC331117 
Bpl55 
GAACTAGCATGGCAGCTAGATCTAG 
GAACCATCAGAACAACTCAACCTGC 
(ATC)13 
150 
KC331119 
Bp221 
TATTGCTGAGCCTGCTTGGCAG 
GCCAAGCTGCTGCAGTCCCAGCAGG 
(GTT)19 
210 
KC331123 
Bp230 
CCAAGGACACATGCCCATATGTCTC 
GTCTTCTGAATGGACCGAGTCTGGC 
(TCA)16 
150 
KC331124 
Bp239 
CAATTTGTGAATCTCCAGGTCAG 
GAACGCCAATCACAGCCTGGCAGTG 
(CAA)19 
160 
KC331127 
Bp275 
TTACGCGCCTGGTGTAGCCTAGCTC 
CCTACGTGCGGCTCACATGTCTGG 
(ATCAGA)9 
170 
KC331130 
Bp301 
CATTATAGCTACAGAGCGGCTGG 
CTGAGTGGGGTTCACCGCAGTCTG 
(GATA)12 
150 
KC331134 
Bp473 
CGGCACTCCGTGAACACAGCGG 
ATGTGCTCTTTGGCTGCAG 
(CACT)8 
170 
KC331144 
Bp489 
AGCGAGGTGAATGGTTCAATTCCAG 
GCAAAACAATTGCCCATACTGGGAC 
(TAG) 10 
250 
KC331145 
Bp501 
ACTCTCGCTGTGTTATTGACAGCGC 
TCAGTTGACTGTGATCTCATGCGAC 
(ATT/GTT)15 
150 
KC331146 
Bp531 
CAGGTCTTTGTGACATGC C AAGCTG 
CTGACTGCTTGAGGGATCGCAGCAG 
(ATG/GTG)16 
140 
KC331148 
cases, the 2-base alleles reflect variation in the untar- 
geted sequence adjacent to the targeted repeat motif. 
All alleles at these loci were nevertheless scored un- 
equivocally through the use of Beckman Coulter frag- 
ment analysis software, and they were treated as di- 
nucleotide repeats in downstream analyses. 
In all 4 samples of Gulf Menhaden, 3 of 19 loci re- 
vealed evidence of null alleles (Table 2). These loci were 
BP017 (overall null allele frequency p n =0.17), BP301 
(overall null p n =0.25), and AF039660 (overall null 
p n =0.30). The loci BP301 and AF039660 also showed 
evidence of genotyping errors due to excessive stutter 
in multiple populations. Four remaining loci revealed 
evidence of null alleles in at least 2 populations: BP155 
(overall null p n = 0.05), BP239 (overall null p n =0.03), 
BP275 (overall null p n =0.06), and AF039661 (overall 
null p n =0.10). A final 2 loci (AF039658 and AF049462) 
revealed evidence of null alleles at very low frequency 
(p n <0.05) in single populations. 
The locus BP155 failed to amplify in samples of 
Atlantic Menhaden, Finescale Menhaden, or Yellow- 
fin Menhaden. There was evidence for null alleles in 
Atlantic Menhaden at the loci BP017, BP230, BP239, 
BP301, AF039658, AF0399660, and AF049462 (Table 
3). In Finescale Menhaden, 2 loci showed evidence of 
null alleles (BP301, AF039660). In Yellowfin Menha- 
den, there was evidence of null alleles at only a single 
locus, BP301. 
Within Gulf Menhaden, there was no evidence for 
linkage disequilibrium between any pair of loci. Across 
all loci, there was a range of 6 observed alleles (BP489, 
AF039661) to 26 observed alleles (AF049462), with a 
mean of 14.8 alleles per locus (Table 2). When loci that 
had null alleles were included, there was an average 
observed heterozygosity of 77 0 = 0.64, and an average 
expected heterozygosity of 77 e =0.74. With null loci re- 
moved, both observed and expected heterozygosity av- 
eraged 77=0.71. Eight loci deviated significantly from 
Hardy-Weinberg expected genotypes before Bonferroni 
adjustment. Each of these disequilibrium loci indicated 
evidence of null alleles. After Bonferroni adjustment, 
only 4 loci significantly deviated from HWE. These 4 
disequilibrium loci (BP017, BP301, and AF039660, 
AF039661) also had the highest estimated null allele 
frequency among all loci examined (p n >0.10 in each 
case). 
Within other species of Brevoortia, overall genetic 
variability was greatly reduced, in comparison to Gulf 
Menhaden, for the genetic loci examined. The mean ex- 
pected heterozygosity was 77 e =0.63 in Atlantic Menha- 
den, T7 e =0.37 in Yellowfin Menhaden, and 77 e =0.44 in 
Finescale Menhaden. Heterozygosity across all loci was 
significantly higher in Gulf Menhaden than in the 3 
Brevoortia congeners on the basis of the Wilcoxon test 
(P<0.001, in each case). The mean PIC values for At- 
lantic Menhaden, Yellowfin Menhaden, and Finescale 
Menhaden were 0.617, 0.356, and 0.429, respectively 
(compared with a PIC value of 0.734 for Gulf Menha- 
den). The PIC value across loci was significantly higher 
in Gulf Menhaden than in any of the other 3 Brevoor- 
tia species on the basis of the Wilcoxon test (P<0.001, 
in each case). 
