Low mitochondrial DNA variation among American alligators and a novel non‐coding region in crocodilians

Abstract

We analyzed 1317-1823 base pairs (bp) of mitochondrial DNA sequence beginning in the 5' end of cytochrome b (cyt b) and ending in the central domain of the control region for 25 American alligators (Alligator mississippiensis) and compared these to a homologous sequence from a Chinese alligator (A. sinensis). Both species share a non-coding spacer between cyt b and tRNA(Thr). Chinese alligator cyt b differs from that of the American alligator by 17.5% at the nucleotide level and 13.8% for inferred amino acids, which is consistent with their presumed ancient divergence. Only two cyt b haplotypes were detected among the 25 American alligators (693-1199 bp surveyed), with one haplotype shared among 24 individuals. One alligator from Mississippi differed from all other alligators by a single silent substitution. The control region contained only slightly more variation among the 25 American alligators, with two variable positions (624 bp surveyed), yielding three haplotypes with 22, two, and one individuals in each of these groups. Previous genetic studies examining allozymes and the proportion of variable microsatellite DNA loci also found low levels of genetic diversity in American alligators. However, in contrast with allozymes, microsatellites, and morphology, the mtDNA data shows no evidence of differentiation among populations from the extremes of the species range. These results suggest that American alligators underwent a severe population bottleneck in the late Pleistocene, resulting in nearly homogenous mtDNA among all American alligators today.

Mycoplasma alligatorisInfection Promotes CD95 (FasR) Expression and Apoptosis of Primary Cardiac Fibroblasts

“…The cells were passaged without attempts to transform or immortalize them, in order to retain the in vivo differentiated characteristics of the cell types (11,12). The source species of each cell line was confirmed by mitochondrial DNA genotyping (26).…”

Genetic characterization of captive Cuban crocodiles (Crocodylus rhombifer) and evidence of hybridization with the American crocodile (Crocodylus acutus)

“…Primers drL15459 (modified from Glenn et al, 2002) and CR2HA (modified from Ray and Densmore, 2002) were used to amplify the tRNA Pro -tRNA phe -D-loop region (442 bp; Table 1) for only C. rhombifer and C. acutus. Polymerase chain reactions (PCR) were performed in 50 \ih volumes using 0.50 (iL of total genomic DNA (tDNA) (50ng/nL), 36.25 nL of ddH 2 0, 10 nL of buffer (0.3 M TRIS, 0.0175 M MgClg, and 0.075 M (NH 4 ) 2 S0 4 ), 2.0 \xL of 2.5 mM dNTPs, 0.50 nL (10 mM) of forward primer, 0.50 (iL (10 mM) reverse of primer and 0.25 (iL (1.25 U) of Promega Taq polymerase (Promega Corp., Madison, WI).…”

Population genetic analysis of Caiman crocodilus (Linnaeus, 1758) from South America

“…The mitochondrial cytochrome Vasconcelos et al 221 b gene was amplified via Polymerase Chain Reaction (PCR) using the primers L14254 (5'-ATGACCCACCAACTACG AAAAT-3') from Glenn et al (2002) and H15982 (5'-TCC CTRGCTTTGGTAGCCAGG-3') from Farias et al (2004). PCR reactions were carried out in a final volume of 25 µL and contained 11.7 µL of ddH 2 O, 3 µL of MgCl 2 (25mM), 2.5 µL of dNTPs (10 mM), 2.5 µL of 10x buffer (100 mM Tris-HCl, 500 mM KCl), 2 µL of each primer (2 µM), 0.3 µL of Taq DNA Polymerase (5 U/µL) and 1 µL of DNA (concentration varied between 50 ng and 100 ng).…”

“…Each reaction contained 4 µL of amplified DNA product (~30 ng), 2 µL of primer (L14254 for the 5' segment of the amplified DNA fragment, and L14731 (5'-TCGTGCCAT GAATTTGAG-3') from Glenn et al (2002) as an internal primer for the 3' portion of our DNA fragment), 2 µL of 5x replacement buffer (400 mM Tris-HCl pH 9.0, 10 mM MgCl 2 ) and 2 µL of DYEnamic ET Dye Terminator mix (Amersham Bioscience, São Paulo). Cycle sequencing PCR conditions were as follows: denaturation at 93°C for 15 s, primer annealing at 50°C for 35 s, and primer extension at 60°C for 120 s; these three steps were repeated 35 times.…”