LEXEN: Longevity and Diversity of Microorganisms Entrapped in Tropical and Polar Ice Cores

该项目旨在确定从青藏高原、南美安第斯山脉和两极地区等不同环境中采集的冰芯中可以复苏的微生物的种类、丰度和古老性。将对PCR扩增的小亚基（16 S和18 S）rDNA进行克隆和测序，以确定这些冰核中存在的或曾经存在的对实验室培养难以抵抗的冰核类型的范围。 这些实验在概念上非常简单，在技术上只需要常规的微生物和分子生物学程序;然而，防止微生物和外源核酸的污染将是至关重要的。 因此，将专门建造一个自动冰取样装置来消除这个问题。 冰芯在严格控制的条件下在BPRC存档，并且大多数地点都有几个冰芯，因此可以进行多次采样以评估不同的程序并确认积极的结果，而无需实地考察。 拟议的实验将确定在不同环境条件下被困在冰中的微生物的寿命，沉积在同一地理位置但相隔数千年或可能数十万年的微生物的多样性，以及沉积在同一考古时期但地理位置非常不同和偏远的微生物的多样性。 这可能是第一次将PCR技术应用于从非常古老的冰芯中扩增材料的共同努力，而且似乎很有可能“灭绝”基因，或者至少是灭绝基因的片段将被扩增，从而促进与其当代同源物序列的有趣比较。 虽然许多核心分离株可能是放线菌，但评价核心分离株的新型抗生素合成超出了本提案的探索性目标。 然而，在抗生素广泛使用之前，确定被困在冰中的微生物的抗生素耐药性模式将是常规的。

This project is to determine the phylotypes, abundance and antiquity of microorganisms that can be resuscitated from within ice cores taken from such diverse environments as the Tibetan Plateau, the South American Andes, and both polar regions. PCR-amplified small subunit (16S and 18S) rDNAs will be cloned and sequenced to determine the range of phylotypes that are, or were, present in these ice cores that are refractory to laboratory culture. The experiments are conceptually very simple, and technically require only routine microbiological and molecular biology procedures; however, preventing contamination with microorganisms and nucleic acids from exogenous sources will be critical. An automated ice-sampling device will therefore be constructed specifically to eliminate this problem. The ice cores are archived under carefully controlled conditions at the BPRC, and several cores area available from most locations, so that multiple samplings to evaluate different procedures and to confirm positive results are possible without field trips. The experiments proposed will determine the longevity of microorganisms entrapped in ice under different environmental conditions, the diversity of microorganisms deposited at the same geographical sites, but separated by thousands, or possibly by hundreds of thousands of years, and the diversity of microorganisms deposited at the same archeological times but at geographically very different and remote locations. This might be the first concerted effort to apply PCR technology to amplify material from within very ancient ice cores, and it seems very likely that "extinct" genes, or at least fragments of extinct genes will be amplified, facilitating intriguing comparisons with the sequences of their contemporary homologs. Although many of the core isolates are likely to be Actinomycetes, evaluating novel antibiotic synthesis by core isolates is beyond the exploratory goals of this proposal. Determining the antibiotic resistance patterns of microorgan isms that were entrapped in ice before the widespread use of antibiotics will, however, be routinely undertaken.