Cloning Of D-Loop And HIF-1 Alpha Gene From Vertebrates In Qing-zang Plateau

Gymnocypris przewalskii, also named scaleless carp, is the only economical aquatic in lake Qinghai. There has been few previous molecular work of population genetics structure about this species. The author used mtDNA D-Loop sequence as genetic marker to resolve the population genetic structure of scaleless carp in lake Qinghai. Partial sequences (639bp) of mtDNA D-Loop of Gymnocypris przewalskii przewalskii and Gymnocypris eckloni have been amplified and sequenced successfully (accession numbers: AY850397 and AY850398). 9 haplotypes have been identified among 60 scaleless carps with SSCP. Base composition and variance have been analyzed. Nucleotide diversity (π), haplotype diversity (h), Fst and Nm have been calculated. The phylogenetic trees constructed by multiple methods (NJ and MP) supported almost the same topology. Our molecular results showed that D-Loop diversity in scaleless carp was higher, but there was no significant differentiation (P>0.1) in results of Tajima's D and Fu and Li's D statistic. Also there was no significant genetic structure differentiation among populations of Heima river, Buha river, Quanji river, Shaliu river and lake Qinghai according to the result of Fst analysis. There were high gene flows among different populations of scaleless carp according to the result of Nm calculation.Hypoxia-inducible factor 1 is a transcription factor composed of HIF-1α and HIF-1β. It is an essential mediator of oxygen homeostasis. Root vole (Microtus oeconomus) and Tibetan antelope (Pantholops hodgsoni) are two native mammals at Qinghai-Tibetan plateau, and well acclimatized to environmental hypoxia. In this study, we have cloned cDNA encoding HIF-1α from the liver of Root vole and the muscle of Tibetan antelope (accession numbers: DQ010149 and AY971808), and examined its expression of mRNA and protein in heart, brain, liver, kidney, lung and muscle of Root vole. The deduced HIF-1α protein sequence (823 amino acids) of Tibetan antelope showed 95%, 90%, 90%, 92%, 92%, 99%, 99%, 79%, 57% and 55%identities with those of the human, rat, mouse, vole, pika, bovine, yak, chicken, scale less carp and rainbow trout, respectively. Meanwhile, the deduced HIF-1α protein sequence (828 amino acids) of Root vole showed 91%, 94%, 93%, 90%, 92%, 92%, 78%, 58% and 55% identities with those of the human, rat, mouse, pika, bovine, yak, chicken, scaleless carp and rainbow trout. Northern blot analyses showed that HIF-1α mRNA in the muscle of Root vole was expressed weaker than other five tissues. Western blot analyses showed that HIF-1α protein was expressed most abundantly in the kidney and liver, followed by other four tissues. Our results suggest that HIF-1α is very conservative among the animals and may play an important role in adaptation to hypoxia of Root vole and Tibetan antelope.