| There was long cultivation history in our country of Phaseolus vulgaris L. Homonym andsynonym were common phenomena on the market, not only to put breeding researchers out, butalso to do cause huge damage in peasant production.The seeds of seventeen common bean in different recovery years and parts were taken foranalysis the seed morphology and field planting and the seeds storage proteins with (SDS-PAGE,Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis) electrophoresis and (RAPD,Random Amplified Polymorphic DNA) molecular marker technology, to find a quickly andsteadily method to Identify Seeds Authenticity and Purity of Kidney Bean. Thus, it couldprovide technical reference for kidney bean breeding and department of producing cotton stockseeds.The seeds of seventeen common beans in2011and2012reserving years were taken forobservation and analysis the seeds morphology, we observed and measured patterns, length,width, thickness, shape and100-seed weight of the seeds. The results show that the seventeencommon beans were divided into six groups, No.1~4varieties which were black bluearabesquitic over creamy white and variance analysis at5%significant level with length, width,thickness and100-seed weight of the seeds had no significant difference. No.5and6varietieswhich were reseda arabesquitic over milk white and there were no significant difference oflength, width, thickness and100-seed weight of the seeds. No.7~11varieties which wereamaranth arabesquitic over milk white and variance analysis at1%significant level with length,width, thickness and100-seed weight of the seeds had no significant difference. No.12and13varieties which were reseda arabesquitic over cream-colored and there were no significantdifference of length, width, thickness and100-seed weight of the seeds. No.14and15varietieswhich were puce arabesquitic over brown and there were no significant difference of length,width, thickness and100-seed weight of the seeds. No.16and17varieties which were Creamywhite and there were no significant difference of length, width, thickness and100-seed weightof the seeds. Because of the seeds morphology in every group were very similar, we couldn’tdistinguish them by morphological observation. Therefore, we could identify accurately andquickly by field planting and SDS-PAGE electrophoresis and RAPD molecular marker.The method of field planting was used in this experiment, the seeds of seventeen commonbeans which were similar in shape in2011and2012reserving years were planted in the open field. They were growed in the same environment. To make sure the difference and authenticitybetween different varieties of kidney bean via the morphological characteristics of seedlingstage, pod and yield. Compare to2011and2012in every groups of every variety,the appropriateidentification index of the first group were corolla color, shape of pod, the position of beak ofpod, essential colour of tender pod, curvature of tender pod; the appropriate identification indexof the second group were hypocotyl color, shape of pod, the position of beak of pod, curvatureof tender pod; the appropriate identification index of the third group were hypocotyl color,corolla color, shape of pod, the position of beak of pod, the position of beak of pod, curvature oftender pod; the appropriate identification index of fourth group were shape of pod, the positionof beak of pod; the appropriate identification index of fifth group were shape of pod, theposition of beak of pod, without joining yarn, the joining yarn colour of pod. Compare to kidneybeans between year2011and2012, the early flowering season was from June27thto July5thin2013, and acre yield was increased in year2012and every variety was add25kg on average.The results shown that there were significant difference between different varieties of kidneybeans. The acre yield of2012reserving year of kidney bean were higher than2011reservingyear, because of the longer storage time, the lower growth potential of seeds. According toprocedure of inspection crop seeds, the experiments examined the purity of variety of kidneybean by field planting. It revealed that field estimated value70.6percentage of2012reservingyear purity of variety exceed92.8percentage, there were only five varieties which were No.9,12,13,16and17less than92.8percentage. Therefore, the field planting method could use toidentify the authenticity and purity of kidney bean. And we need a novel method to overcomethe long growth cycle and made it ignore environmental implication.The seeds of seventeen common bean which were similar in shape of storage protein in2011and2012reserving years were used in SDS-PAGE electrophoresis, the unique bands A、B、C、D、E、F、G and H were found, and the software of Bandscan5.0were used to analyze thedifference of expressing quantity and the percentage account for the total protein of the lane ofevery bands. Protein electrophoresis of kidney bean seed in year2011, the result shown that A、B、D、E、F were characteristic bands of No.1~4varieties, and B、C、F、G were characteristicbands of No.7~11varieties, B、D、E、F、H were characteristic bands of No.12and13varieties,C、D were characteristic bands of No.14and15varieties, B、F、G were characteristic bands ofNo.16and17varieties. Protein electrophoresis of kidney bean seed in year2012, the resultshown that C was characteristic band of No.1~4varieties, B、C、G were characteristic bands ofNo.7~11varieties，C was characteristic band of No.12and13varieties, B、H were characteristicbands of No.14and15varieties. Protein electrophoresis of kidney bean leaves in year2011, the result shown that B、F were characteristic bands of No.1~4varieties, C、F、G、H werecharacteristic bands of No.7~11varieties, B、C、D、E were characteristic bands of No.12and13varieties, C was characteristic band of No.14and15varieties, Protein electrophoresis of kidneybean leaves in year2012, the result shown that B、F were characteristic bands of No.1~4varieties, C、F、G、H were characteristic bands of No.7~11varieties，B、C、D、E werecharacteristic bands of No.12and13varieties, C was characteristic band of No.14and15varieties. The authenticity of other varieties could be identified by analyzing the difference ofprotein expression quantity. The protein zones comparative of year2011and2012, the materialsby year2012presented the characteristic bands of C、E、H and these as year2011weredisappeared, long time storage could result the degradation of protein. It revealed that the wholetest material were different variety and truth. The comparison of kidney bean between7and8varieties, the result showed that the seven varieties was mixed in the eight varieties for Fappeared in the eight varieties. The F and G lanes expressions of six groups were significantlyincreased at grain number level of No.7, and the bands were gradually darkening than before.The experiment could be done only use a grain of seed of kidney bean even though test materialwas shortage. Identification of seeds authenticity and purity in kidney bean by SDS-PAGEelectrophoresis is the first time in the domestic, it was suitable for identification of seedsauthenticity and purity in kidney bean for good stability and replication and with a significantresolution ratio of detection.The seeds of seventeen common bean which were similar in shape in2012reserving yearswhich the true leaf was appear in fifteenth day were taken for RAPD molecular markerelectrophoresis. The result shown that they were all genomic DNA and there was no RNAimpurity in it. Twenty four primers were screened,5selected primers produced clearpolymorphic patterns.15specificity RAPD bands which were1500bp,875bp,750bp,700bp,690bp,625bp,560bp,500bp,490bp,440bp,420bp,375bp,330bp,250bp and240bp wereamplified of kidney bean in2012year, the length of amplification specificity fragments between1500bp and240bp. No.1~4, No.7~11, No.16and No.17varieties were identified by primer one,and bands were obvious difference.375bp and1500bp were characteristic bands of No.1~4varieties,420bp、500bp and750bp were characteristic bands of No.7~11varieties,375bp and750bp were characteristic bands of No.16and17varieties. No.1, No.4, No.5, No.6, No.7, No.9,No.11~15varieties were identified by primer two.330bp、375bp and490bp were characteristicbands of No.1~4varieties,240bp、250bp and500bp were characteristic bands of No.7~11varieties,250bp and500bp were characteristic bands of No.12and13varieties. No.3.No.1~4,No.9and No.13varieties were identified by primer three,375bp、440bp and875bp were characteristic bands of No.1~4varieties,700bp was characteristic band of No.7~11varieties,440bp and500bp were characteristic bands of No.12and13varieties..No.1~4, No.9~17varieties were identified by primer three,375bp and875bp were characteristic bands of No.1~4varieties,375bp、500bp and625bp was characteristic band of No.7~11varieties,875bp wascharacteristic band of No.14and15varieties,625bp and875bp were characteristic bands ofNo.16and17varieties. The result of primer five amplification of No.1~6and No.9~17varietiesshown that it was clear and good,440bp、560bp、625bp and875bp were characteristic bands ofNo.1~4varieties,440bp and875bp were characteristic bands of No.7~11varieties,440bp wascharacteristic band of No.12and13varieties,690bp was characteristic band of No.16and17varieties. It could be distinguished by different shades and quantity of bands in everygroup.No.5and6varieties in year2012were mixed by human, the purity spectrum of RAPD ofNo.5and No.6was good and the purity was94.12%by year2012. |
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