| Gossypium raimondii is one of the spices in diploid genome D5, the smallest genome(880Mb) in Gossypium, and it is widely known the D-subgenome donor of Allotetraploid. There is a potential resistance against drought, verticillium wilt, bollworm, leafhoppers, thrips, leaf insects in G. raimondii, and also potential to increase yield, fiber quality in cotton breeding. Genome sequencing of G. raimondiih, has been completed, which may lead to disclosing its rich mining resources of wild-type characteristic.Gossypium darwinii is tetraploid(AD)5, mainly distributed in the Galapagos Islands, surviving the wild harsh environment, which implying the potential for drought, hot wind, salinity resistance. Salinity trait for G. darwinii plays a significant role breeding research.CCICR(Chinese Central Institute of Cotton Research) is the latest identified cotton endemic transposon family. There is a huge proportion difference in the cotton genome A and D genome species sequence, 36.2% in the A genome sequence, but in D genome proportion is almost zero, which is considered the main cause of size differences beyween A and D genome. The BAC library has been screened by to detect CCICR specific monoclone for further analyzing the evolution of the distribution in the cotton genus.Based on the existing construction methods for diploid and tetraploid BAC libraries, we successfully constructed wild cotton G. raimondii and G. darwinii cotton BAC library. With existing SSR molecular marker on CCICR primers NAU1201 we screened the G. barbadense pima90-53, G. darwinii and G. tomentosum to obtain single BAC clone containing the CCICR. The results as follows:1. Using the p Indigo BAC-5(Hind III) and p HZAUBAC1 as carrier, G. raimondii and G. darwinii BAC library were constructed. G. raimondii BAC library contains 26,880 clones with an average insert size of 127 kb, covering about 3.7 times the cotton genome, cloning load rate less than 5%. Gossypium darwinii BAC library have 200,832 clones covering G. darwinii genome 10 times. The average insert sequence length is 122 kb, cloning load rate less than 5%. After testing, BAC monoclonal can be stably inherited more than 125 generations,The stability of the library meet the requirements for follow-up experiment. The construction of wild cotton BAC library is of great significance in development and utilization for resources of wild cotton2. After three-stage PCR screening of the G. barbadense Pima90-53, G. darwinii and G. tomentosum BAC libraries with the genetic linkage group SSR primers NAU1201, thirty-three single BAC clones containing CCICR were obtained. BAC clones 31-G-32, 37-F-10, 42-C-10, 170-G-11, 170-G-12, 170-G-13, 242-C-11, 299-I-16, 299-N-20, 317-K-22 and 317-L-22 are from G. barbadense Pima90-53. BAC clones 193-F-24, 193-G-24, 193-H-24, 234-I-9, 302-M-13, 325-P-20, 395-D-6, 395-E-5, 408-J-6, 413-L-18 and 433-A-20 are from G. darwinii. BAC clones 30-B-3, 30-B-4, 6-O-16, 6-O-17, 89-J-11, 91-K-2, 91-K-3 and 91-K-4 are from G. tomentosum. The screening of these three libraries in three CCICR cotton BAC library lay a good foundation for co-linear analysis and further analysis of the evolution of the distribution in the cotton. |
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