佛坪三官宙庙地区大熊猫种群数量的DNA指纹分析

应用同位素标记大熊猫基因指纹探针Ｆ２ＺＧＰ９６０６８０８０１，以秦岭南坡中段佛坪国家级大熊猫自然保护区三官庙范围内采集的大熊猫粪便样品作材料，进行了ＤＮＡ作指纹检测，（１）在相同或不同时间，领域采粪便样品，呈出现相同或不同的ＤＮＡ指纹图谱，达到个体认定的目的，进一步表明了大熊猫的尽量新鲜的粪便，可以作为ＤＮＡ指纹分析材料，进行野生生种群数量调查；（２）根据检测２１个粪便样品的结果，无误地定了三官庙     

利用LZF－1DNA指纹探针进行家系的父权鉴定

ＬＺＦ－１ＤＮＡ指纹探针经同位素γ－３２Ｐ－ＡＴＰ标记后，检测了４个家系１３个个体血样的ＤＮＡ指纹．结果表明，父母的遗传物质在子代中的传递符合孟德尔遗传规律，无误地确定了４个家系中的亲子血缘关系．ＬＺＦ－１ＤＮＡ指纹探针在亲权鉴定中的父权概率是０．９９９６４，达到了父权认定的目的     

利用LZF—1DNA指纹探针进行家系的父权鉴定

ＬＺＦ－１ＤＮＡ指纹探针经同位素γ＾３２Ｐ－ＡＴＰ标记后，检测了４个家系１３个体血样的ＤＮＡ指纹，结果表明，父母的遗传物质在子代中的传递符合孟德尔遗传规律，无误地确定了４个家系中的亲子血缘关系，ＬＺＦ－１ＤＮＡ指纹针在亲权鉴定中的父权概率是０．９９９６４，达到了父权认定的目的。     

大熊猫肌肉氨基酸含量的初步研究

测定了３只大熊猫的头部、背部和腿部等１０个肌肉样品中１７种氨基酸（ＡＡ）含量,并分析了各样品ＡＡ与Ｌｙｓ含量的比值．结果表明：对同一只大熊猫,不同部位的肌肉样品中各ＡＡ含量无明显差异;对同一个肌肉样品,１７种ＡＡ含量比较,Ｇｌｕ含量最高,Ａｓｐ、Ａｒｇ、Ｌｙｓ、Ｌｅｕ、Ａｌａ、Ｇｌｙ、Ｖａｌ、Ｐｒｏ、Ｔｈｒ、Ｐｈｅ等含量较高;不同年龄大熊猫肌肉ＡＡ含量有差异,除极少数ＡＡ外,未成熟雄性大熊猫肌肉ＡＡ含量高于成熟和老龄雌性大熊猫,且有显著性差异（Ｐ＜０．０５或Ｐ＜０．０１）;成熟雌性大熊猫肌肉中,除Ｐｒｏ外,其余所有ＡＡ含量与其年龄无显著差异（Ｐ＞０．０５）;除极少数ＡＡ外,不同年龄、性别大熊猫肌肉样品中各ＡＡ与Ｌｙｓ含量的比值无明显差异     

寡核苷酸探针LZF－I进行两栖类DNA指纹分析的初步研究

利用寡核苷酸探针ＬＺＦ－Ｉ对两栖类的３属８种２８只个体的冷冻和甲醛固定的肝组织作了检测．所获ＤＮＡ指纹图在个体、种和属间具有特异性．表明该探针进行ＤＮＡ指纹检测可为两栖动物种、属分类提供依据．     

游蛇科（Colubridae）10种蛇的随机扩增多态DNA研究

用随机扩增多态ＤＮＡ（ＲＡＰＤ）技术对游蛇科１０种蛇基因组ＤＮＡ的多态性进行了分子遗传标记的研究．所得数据经聚类分析结果提示：１．蛇类种内个体间存在着遗传多样性即个体间的差异，随机扩增多态ＤＮＡ片段共享度的分析表明，种间差异显著大于种内个体间的差异，说明在研究种间系统演化关系时．可以用随机取样的个体代表一个种作种间比较．２．锦蛇属是一高度分化的大属，本研究中的５种锦蛇间．玉斑锦蛇和红点锦蛇关系较近．可以井为１组．另３种归为１组还是分为３个不同的组尚难定论．３．游蛇科６个属之间．锦蛇属、乌梢蛇属和鼠蛇属３属间亲缘关系较近．Ｒｈａｂｏｐｈｉｓ和Ｓｉｎｏｎａｔｒｉｘ与链蛇属较近．它们可能代表该科中较原始的类群．     

用同位素标记LZF—1 DNA指纹探针进行人的DNA指纹分析

同位素γ－32P－ATP对LZF－1DNA指纹探针作5＇末端标记后，检测成都地区人群中84名随机个体的DNA指纹，获得了清晰易辨的具有高度个体特异性的DNA指纹图谱．谱带平均数为17．667条，平均等位基因频率为0．167，探针的鉴别机率为4．862×10－10．表明LZF－1DNA指纹探针完全达到了法医学检验中鉴别个体的目的．     

玉米种子的DNA指纹计算机化鉴定

对１２个优良玉米自效系的ＤＮＡ进行了ＲＡＰＤ分析，共筛选了２５０个Ｏｐｅｒｏｎ引物，其中１２个引物共扩增出５９个多态性产物，根据这５９个多态性ＲＡＰＤ产物，对１２个自交系进行了聚类分析，把它们分为３个群，结果与根据系谱法的聚类结果一致。经过优选，用ＯＰＮ－１１扩增出的１１种ＤＮＡ带，建立了这１２个自交系的ＲＡＰＤ－ＤＮＡ指纹图谱。在该图谱呼弱个自交系都具有特异的ＤＮＡ指纹，可以与其它自效纱相区别。     

寡核苷酸探针LZP—I进行两栖类DNA指纹分析的初步研究

利用寡核苷酸探针ＬＺＦ－Ｉ对两栖类的３属８种２８只个体的冷冻和甲醋固定的肝组织作了检测，所获ＤＮＡ指纹图在个体，种和属间具有特异性，表明该针进行ＤＮＡ指纹检测可为两栖动物种，属分类提供依据。     

大熊猫基因组微卫生DNA的分离与序列分析

以pBS^ 为克隆载体,构建圈养大熊猫基因组DNA的小插入文库，分别用生物素标记的（CAC）5和γ-^32P-dATP标记的（CA）15寡核苷酸为探针，对重组质粒进行斑点杂交，获得15个阳性克隆，并测定了插入片段的DNA序列，其中有一部分序列已被分析，本文对剩余的8个DNA序列进行分析，发现它们大小不一，从259bp-881bp，有6个序列含有不同重复单位的一、二、四核苷酸的完整串联重复，斑点杂交和Southern杂交证明，这些序列均来自大熊猫基因组，上述结果为利用PCR技术研究大熊猫遗传多样性和了解大熊猫基因组结构积累资料。     

野化培训大熊猫采食和人为砍伐对拐棍竹无性系种群生物量的影响

采用固定样方法和定位观察法连续测定了2003～2007年间卧龙自然保护区大熊猫野化培训圈内及其附近区域的拐棍竹无性系种群数量和生长发育特性等参数,运用收获法与非破坏性重量估测法建立了不同龄级和残桩的竹子种群和分株生物量估测模型,进而利用最佳模型计算并评估了野化培训大熊猫采食和人为砍伐对拐棍竹无性系种群生物量和植株个体生物量的影响.结果表明:在环境条件、种群密度、生长发育特征和种群生物量等基本相似的基础上,大熊猫采食和人为砍伐不仅降低实验期间的竹子生物生产力,而且影响到后期阶段实验种群的恢复与发展.大熊猫采食样方中的竹子种群生物量虽然较对照样方低,就竹笋生物量而言,约为对照的57.79%,这与野化培训圈的面积较小、竹种单一而使采食比重(67.07%)较大有关,但其各龄级植株个体生物量均能达到大熊猫的取食利用标准(仅2004年生竹除外),具有持续供给大熊猫食物资源的潜力;而人为砍伐措施与大熊猫采食相比,影响效果极为强烈,它严重降低了拐棍竹无性系的种群生物量,尤其是竹笋重量更是如此,仅为对照样方的14.69%,且植株个体鲜重远低于大熊猫的觅食条件.因此,竹笋和无性系植株的生物量是大熊猫采食标准的主要因素.     

Conservation Implications of Drastic Reductions in the Smallest and Most Isolated Populations of Giant Pandas

Abstract: In conservation biology, understanding the causes of endangerment is a key step to devising effective conservation strategies. We used molecular evidence (coalescent simulations of population changes from microsatellite data) and historical information (habitat and human population changes) to investigate how the most‐isolated populations of giant pandas (Ailuropoda melanoleuca) in the Xiaoxiangling Mountains became highly endangered. These populations experienced a strong, recent demographic reduction (60‐fold), starting approximately 250 years BP. Explosion of the human population and use of non‐native crop species at the peak of the Qing Empire resulted in land‐use changes, deforestation, and habitat fragmentation, which are likely to have led to the drastic reduction of the most‐isolated populations of giant pandas. We predict that demographic, genetic, and environmental factors will lead to extinction of giant pandas in the Xiaoxiangling Mountains in the future if the population remains isolated. Therefore, a targeted conservation action—translocation—has been proposed and is being implemented by the Chinese goverment.     

Potential solar radiation pattern in relation to the monthly distribution of giant pandas in Foping Nature Reserve, China

Solar radiation is an important parameter in ecological process modeling, hydrological modeling and bio-physical modeling. However, models focusing on solar radiation in relation to giant panda habitat and seasonal distribution are limited. The research aims to form spatial models of 12 month solar radiation patterns and to investigate the relation between the solar radiation patterns and the monthly distribution patterns of giant pandas. The solar radiation model of Kumar et al. was adopted for this study in Foping Nature Reserve (NR), China. By comparing twelve monthly solar radiation patterns and calculating statistics such as maximum, minimum, mean and standard deviation of the solar radiation, diversified solar radiation patterns over different months were obtained. Maximum solar radiation occurred in June and July, while minimum solar radiation occurred in December and January. The annual sum of solar radiation was 6954 MJ/m² in Foping NR. The range in solar radiation was smaller in hot months and larger in cold months. Radio tracking data of giant pandas were collected for twelve months and the ensuing maps were overlaid with the twelve-month solar radiation map to analyze the relation between the giant panda's monthly distribution and solar radiation. Our results showed that giant pandas prefer areas with lower solar radiation in warm months and select areas with higher solar radiation in cold months, which illustrates that the distribution of giant pandas is indeed affected by solar radiation. To a certain degree, it also explains the behavior of seasonal movement by giant pandas in Foping NR.     

Patterns of Genetic Diversity in Remaining Giant Panda Populations

Abstract: The giant panda ( Ailuropoda melanoleuca ) is among the more familiar symbols of species conservation. The protection of giant panda populations has been aided recently by the establishment of more and better-managed reserves in existing panda habitat located in six mountain ranges in western China. These remaining populations are becoming increasingly isolated from one another, however, leading to the concern that historic patterns of gene flow will be disrupted and that reduced population sizes will lead to diminished genetic variability. We analyzed four categories of molecular genetic markers (mtDNA restriction-fragment-length polymorphisms [RFLP], mtDNA control region sequences, nuclear multilocus DNA fingerprints, and microsatellite size variation) in giant pandas from three mountain populations (Qionglai, Minshan, and Qinling) to assess current levels of genetic diversity and to detect evidence of historic population subdivisions. The three populations had moderate levels of genetic diversity compared with similarly studied carnivores for all four gene measures, with a slight but consistent reduction in variability apparent in the smaller Qinling population. That population also showed significant differentiation consistent with its isolation since historic times. From a strictly genetic perspective, the giant panda species and the three populations look promising insofar as they have retained a large amount of genetic diversity in each population, although evidence of recent population reduction—likely from habitat loss—is apparent. Ecological management to increase habitat, population expansion, and gene flow would seem an effective strategy to stabilize the decline of this endangered species.     

Proposed Conservation Landscape for Giant Pandas in the Minshan Mountains,China

Abstract: The giant panda (Ailuropoda melanoleuca), is one of the world's most endangered species. Habitat loss and fragmentation have reduced its numbers, shrunk its distribution, and separated the population into isolated subpopulations. Such isolated, small populations are in danger of extinction due to random demographic factors and inbreeding. We used least‐cost modeling as a systematic approach to incorporate satellite imagery and data on ecological and behavioral parameters of the giant panda collected during more than 10 years of field research to design a conservation landscape for giant pandas in the Minshan Mountains. We identified 8 core habitats and 4 potential linkages that would link core habitats CH3, CH4, and CH5 with core habitats CH6, CH7, and CH8. Establishing and integrating the identified habitats with existing reserves would create an efficient reserve network for giant panda conservation. The core habitats had an average density of 4.9 pandas/100 km² and contained approximately 76.6% of the giant panda population. About 45% of the core habitat (3245.4 km²) existed outside the current nature reserves network. Total estimated core habitat decreased between 30.4 and 44.5% with the addition of residential areas and road networks factored into the model. A conservation area for giant panda in the Minshan Mountains should aim to ensure habitat retention and connectivity, improve dispersal potential of corridors, and maintain the evolutionary potential of giant pandas in the face of future environmental changes.     

Giant pandas attend to androgen-related variation in male bleats

Although androgen-dependant traits are predicted to signal overall male quality, no study has examined the response of a nonhuman animal to variation in a known acoustic cue to male androgen levels (steroid hormones that are key drivers of male sexual behaviour). Here, we use a single-speaker approach to present male and female giant pandas with re-synthesised male bleats representing callers with high and low androgen levels. Our results revealed that male and female giant pandas had significantly greater-looking responses, spent more time pacing, and were faster to respond to playbacks of bleats simulating high androgen males. When we analysed the sexes separately, a slightly different response pattern was revealed: whereas males and females still had significantly greater-looking responses and were faster to respond to bleats simulating high androgen males, only male giant pandas tended to spend more time pacing. These findings suggest that vocal cues to male androgen levels are functionally relevant to male and female giant pandas during the breeding season, and constitute the first demonstration that a nonhuman animal could be using a vocal signal to assess male hormonal state. We go on to discuss the ecological relevance of signalling androgen levels in this species’ sexual communication and the possible application of our results to conservation breeding.     

用^32P后标记方法测定典型有毒污染物的DNA加合物

＾３２Ｐ后标记方法测定生物样品里的有毒污染物－ＤＮＡ加合物的过程是用小球菌核酸酶和脾核酸酶将ＤＮＡ及加合物酶解成为２‘－脱氧核苷３’－单磷酸，以它为底物，在Ｔ４多核到激酶的催化下，与γ＾３２ＰＡＴＰ进行文笔磷的转移反应，形成带放射磷的２‘－脱氧核苷３’，５‘－二磷酸酯ＯＤＳ－ＴＬＣ吸附薄层板和ＰＥＩ－ＴＬＣ阴离子交换薄层板将标记过的带放射磷的＾＊３’，５‘－二磷酸酯分离分辨，然后用自显影技术制成加     

High degree of polyandry in Apis dorsata queens detected by DNA microsatellite variability

Workers of six colonies of the giant honeybee Apis dorsata from Sabah, Malaysia (five colonies) and Java (one colony) were genotyped using single locus DNA fingerprinting. The colonies from Sabah nested in colony aggregations of 5 and 28 nests respectively on two trees. Three DNA microsatellite loci (A14, A76, A88) with a total of 27 alleles provided sufficient genetic variability to classify the workers into distinct sub-families revealing the degree of polyandry of the queens. Queens mated on average with 30.17 ± 5.98 drones with a range from 19 to 53. The average effective number of matings per queen was 25.56 ± 11.63. In the total sample of 192 workers, 22 individuals were found that were not offspring of the colony's queen. Three of these were potentially drifted offspring workers from genotyped queens of colonies nesting on the same tree.